AMER. ZOOL., 28:945-962 (1988)
The Structure and Permeability of Integument1
HARVEY B. LILLYWHITE
Department of Zoology, University of Florida,
Gainesville, Florida 32611
AND
PAUL F. A. MADERSON
Department of Biology, Brooklyn College,
Brooklyn, New York 11210
SYNOPSIS. The skin is a heterogeneous, multimembrane system with multiple diffusion
pathways and potentially numerous rate-limiting barriers for specific molecules that are
exchanged with the environment. A broad survey of animals indicates that integumentary
coverings are morphologically, biochemically and embryologically diverse but with common themes of adaptation. The evolutionary proliferation of intercellular junctions, fibrous
or mineralized protective barriers, and lipoid waterproofing barriers emphasize the generality of diffusion limitations. However, the regulation of specific exchange processes
such as the flux rates of respiratory gases entails a complex interaction of multiple factors
affecting both diffusion and perfusion limitations. Consideration of specific pathways and
rate limitations for diffusion of various substances suggests that the permeation of skin
by specific molecules can be partially independent of other exchange processes. Our
understanding of regulated permeability is, however, lacking in mechanistic and integrated
analysis in most cases. Comprehensive understanding of the integument as a regulatory
pathway of communication with the environment will require comparative studies of
specific transport pathways, their rate-limiting resistances, and the interactions as well as
individual regulation of transported molecules.
INTRODUCTION
The integument provides physical protection for internal organs and regulates
the exchange of materials between the
organism and its environment. While various other roles are evident in many species,
the integument is primarily a barrier and
transporting surface. For different animals, a wide range of substances potentially
permeate the skin, and numerous factors
affect their rates of permeation. To understand the limitations of diffusion and the
cutaneous regulation of diffusional
exchange, we must consider both morphological and physiological features of integument.
This article considers the structural
organization of skin as it relates to the
movement of various materials between the
environment and the internal milieu. Studies of integumentary morphology are long
standing, and, in recent years, research on
permeability has been advancing rapidly
with increasing sophistication. Rarely,
however, are these two realms of investigation satisfactorily united in context, and
rarely do the approaches of investigators
consider competing demands on the link
between function and structure. Thus, it
is hoped that the focus of this review will
stimulate interest and future study in these
neglected areas.
Recent and excellent reviews of integumentary morphology in the various animal
taxa are numerous, and this information
will not be repeated here (see, for example,
Bereiter-Hahn et al., 1984, 1986). Rather,
common structural themes will be emphasized in relation to transport phenomena
as elucidated from physicochemical and
physiological studies. Discussion will be
limited to water, ions and respiratory gases
as these have received the most attention
from physiologists.
THE GENERALIZED MORPHOLOGY
OF INTEGUMENT
1
From the Symposium on Cutaneous Exchange of
Cases and Ions presented at the Annual Meeting of
the American Society of Zoologists, 27—30 December
1986, at Nashville, Tennessee.
Before considering the permeability of
specific components of integument, it is
945
946
H . B. LlLLYWHITE AND P . F. A. MADERSON
A
B
^-B — ms
—a
—B
a
sg-
OG
IG
b
FIG. 1. Generalized epidermal organization in (A) a turbellarian, (B) a terrestrial insect, (C) an amphibian,
and (D) a squamate reptile just prior to skin shedding. The examples illustrate features of simple and complex
integuments in both invertebrate and vertebrate animals (not drawn to identical scales). The symbols are as
follows: a, alpha keratin; b, basal laminae or basement membrane; B, beta keratin; c, cement; cl, clear layer;
en, endocuticle; ep, epicuticle; ex, exocuticle; gly, glycocalyx; IG, inner epidermal generation; m, mucus; ms,
mesos layer; OG, outer epidermal generation; sc, stratum corneum; se, syncytial epithelium; sg, stratum germinativum; tw, terminal web; w, wax.
important to review some general organizational features. These considerations will
be restricted to triploblastic animals,
because in diploblastic taxa lacking a mesoderm the epithelia are not free to specialize
at a tissue level that is comparable to the
outer epithelia of more advanced forms
(Fig- 1).
The skin of invertebrates is embryologically as well as structurally diverse, reflecting both phylogenetic and ecological factors. Invertebrates typically possess a
monostratified epidermis that is derived
from ectoderm and overlies a basal lamina
that is either amorphous or fibrous (basement membrane). The epidermis of turbellarians is representative of the simplest
level of organization; it is a single layer of
epithelial cells which overlies muscles and
lacks an external cuticle. The epidermis of
certain Platyhelminthes, Nematoda and
pseudocoelomate phyla is syncytial rather
than cellular. This form is restricted, how-
ever, and has no counterpart among higher
invertebrates or vertebrates. In more
advanced invertebrates, the epidermis may
be thickened and protected by secreted
mucus, cuticle or mineralized structures.
The skin of adult vertebrates is a multicellular, stratified epithelium of ectodermal origin overlying fibrous and vascular
mesoderm. The epidermis never forms a
confining exoskeleton strictly comparable
to that of arthropods, molluscs or echinoderms. All types of cells that are present
throughout the epidermis are represented
by precursors in the basal layer, which proliferates cells that are eventually lost from
the animal's surface. In fishes, mitotic
activity is detectable throughout the epidermis although it is most prevalent in the
deeper layers. In tetrapod vertebrates,
mitotic activity is limited to the basal layer.
Keratin is formed by epidermal cells in all
groups including fishes, but it is a prevalent
structural feature only in epidermis of
SKIN STRUCTURE AND PERMEABILITY
947
amniotes. The form of integument may Collagen and other similar structural
include patterned folds or scales in addi- proteins are common components of invertion to appendages that result from local- tebrate cuticle and frequently form asized epidermal and/or dermal cell prolif- sociations with hyaluronic acid, mucopoeration and differentiation (Maderson, lysaccharides and small amounts of liquid.
1972). Cellular renewal of the epidermis is The structure and composition of these
a vertebrate characteristic that maintains structural elements are related to the
integumentary health and function. In mechanical and durability requirements of
contrast, the maintenance of invertebrate the integument. Some proteins undergo
skin involves replacement of cuticle and sclerotization or "tanning" which stiffens
underlying apical cell membranes without the cuticle and causes the protein to become
cellular renewal.
water-insoluble by cross-linking the adjaThe resistance of a given layer of cellular cent chains, making a more resistant and
cutaneous tissue to chemical invasion is less extensible structure. Further hardendirectly related to its water content (Krogh, ing is achieved by mineralization.
1919) as well as ionic and viscous properThe third important modification of
ties. Therefore, limitations of diffusion are cutaneous form is the presence of lipids
related to the compositional features of a either as a superficial covering, as in insects,
tissue in addition to its thickness and area or sequestered in diffuse or localized layers
(e.g., Shick etal., 1979). Additionally, three within the epidermis, as in various vertemorphological trends in the adaptive evo- brates. These lipids comprise a variably
lution of integuments have had profound effective barrier to water diffusion, and they
effects on the permeability of animal sur- affect the movement of other molecules
faces.
across the skin as well.
First, the integuments of probably all
PERMEATION OF WATER, GASES AND
higher animals have been "tightened" by
IONS: GENERAL PRINCIPLES
intercellular junctions which, among other
things, form occluding barriers by restrictThe integument is a complex, heteroing the diffusion of fluids and solutes geneous structure comprising a cascade of
between cells. In vertebrate tissues "tight resistances to movements of molecules
junctions" (—zonulae occludentes) form belt- through multiple membranes in series.
like regions of intimate contact between Considering the multilayered complexity
plasma membranes of adjacent cells. These of this structure, three fundamental quesare generally impermeable to lanthanum tions arise: (1) What are the pathways of
tracer and contribute to the maintenance molecular movement across the skin? (2)
of osmotic and electrical gradients across What steps in the permeation process are
epithelial layers. Tight junctions are gen- rate-limiting? (3) How do various permeant
erally absent from invertebrate epidermis molecules differ with respect to their facilwith the exception of tunicates, in which ity to pass a given structural barrier?
they are leaky. Septate junctions appear to Although the following sections review
substitute for tight junctions in other information that bears on these topics, the
invertebrates where they are also compar- questions remain largely unanswered.
atively leaky, being permeable to tracers Much current knowledge regarding the
such as lanthanum. The epidermis of fresh- permeability properties of individual biowater and terrestrial organisms typically has logical membranes is incomplete or conmore septa than that of marine inverte- troversial, so, clearly, understanding the
brates, which experiences reduced concen- mechanistic diversity of molecular transtration gradients across the epithelial layer. port across integument will require an
A second trend in evolution is the ongoing commitment to rigorous and
strengthening of the epidermis and its imaginative research.
derivative structures by means of fibrous
To the extent that a multicellular integpolymer layers of protein or, in various ument consists of a number of cell meminvertebrates, a chitin-protein complex. branes in series, the rate limiting step(s)
948
H . B. LlLLYWHITE AND P . F. A. MADERSON
for transport of molecules through the skin described by Fick's diffusional equation.
can be the passage across the membrane/ Both general definitions and interpretacytosol or membrane/environment inter- tions of permeability coefficients are based
face, movement in the interior of the mem- upon, or related to, the Fick relationship.
brane, or diffusion in the cutaneous matri- With respect to water movement, two difces between the membranes, neglecting ferent permeability measurements are freunstirred layers at the skin surface (see quently used. The diffusional water perFederand Pinder, 1988). If transport across meability coefficient (Pd) expresses the
the membranes is fast compared with dif- isotopic flux of water through a unit of area
fusion in the cytosol, movement can then per unit of concentration difference of isobe limited by diffusion polarization related topic water. On the other hand, if concento diffusion-limited liquid zones in series tration differences of impermeant solutes
with the membranes (Neumcke, 1971). cause an osmotic flow of water, the osmotic
Intercellular routes of transport are con- flow through a unit of area per unit of
concentration difference of solute describes
sidered in the next section.
Natural membranes occurring within the the osmotic permeability coefficient (Pf). The
skin are mosaic structures containing lipid term filtration coefficient or hydraulic conbilayers in addition to pores or molecular ductivity applies if the driving force for water
channels. While thermal mobility of molec- flow is a differential of hydrostatic presular species in a diffusing system is inversely sure. In contrast to diffusion, bulk osmotic
related to the molecular mass, the relative or hydrodynamic flow involves the vectodiffusivity of molecules through skin rial movement of an assembly of molecules
depends on considerations of solubility and being driven by an imposed potential. Conpartition coefficients as well. Thus, the sequently, the various permeability coeffiextent to which molecular size or lipid sol- cients may differ quantitatively because of
ubility regulates the penetration of mole- the physical nature of the water movement
cules into cutaneous cells depends on the pathway (see Schafer and Andreoli, 1972;
fractional membrane area occupied by Finkelstein, 1984).
channels and the characteristics of the
If water movement occurs by a solubilitychannels (see Finkelstein and Cass, 1968; diffusion mechanism {i.e., diffusion through
Finkelstein, 1984). Additionally, mem- a lipid bilayer in which water is poorly solbrane proteins may facilitate the flux of uble), then it can be shown that bulk and
water or dissolved molecules either by spe- isotopic water flow are equal such that P f /
cific transport or perhaps by forming a low- Pd = 1 (Cass, 1968). On the other hand, if
resistance pathway between the protein and water transport is through channels, P f /P d
lipid. Various membrane proteins that generally exceeds 1, and the ratio increases
serve as channels for specific ions may also with increasing channel radius (neglecting
permit the concomitant permeation of problems of unstirred layers). The greater
water, nonelectrolytes and dissolved gases. value of Pf results from the fact that osmotic
In generalized terms, molecules cross the water transport occurs by laminar or quaprotein-free bilayer by a solubility-diffu- silaminar flow (Mauro, 1957).
sion mechanism, dissolving into the hydroAqueous pores probably comprise the
phobic regions and then diffusing through major route for water transport in certain
the bilayer subject to the boundary con- epithelia including integument of amphibditions that are established. Lipophilic ians and possibly some aquatic invertemolecules are very permeant, whereas brates (Hevesy et ai, 1935; Koefoed-Johnhydrophilic molecules are relatively imper- son and Ussing, 1953). However, in many
meant. The movement of lipophilic mol- cell membranes the number of ion conecules across the bilayer depends on the ducting channels are apparently too few to
degree of packing and thermal mobility of provide a significant pathway for water
the hydrocarbon chains and on the charge movement (Finkelstein, 1984). With few
of polar head groups of phospholipids.
exceptions, the Pf attributable to memThe free diffusion of a molecular species brane channels is calculated to be about
across an interface or barrier can be 1% that of the value for the entire mem-
SKIN STRUCTURE AND PERMEABILITY
brane, and therefore P f /P d approximate
unity. Thus, the bulk of transcellular water
movement in cutaneous tissues appears to
occur by a solubility-diffusion mechanism
involving the lipid bilayer pathway. Even
so, water flux may occur at the lipid/protein interface as well as via channels formed
by proteins, so that water permeability may
be governed largely by the presence of
membrane-spanning proteins and their
interactions with the lipid bilayer. Diffusional flux of water through lipid bilayers
is reduced by orders of magnitude in the
absence of protein (Carruthers and Melchior, 1983).
The permeation mechanism for water in
lipid bilayers is conceivably driven by the
interaction of lipid polar head groups with
water rather than the solubility of water in
the lipid hydrocarbon (Carruthers and
Melchior, 1983). In any event, water diffuses (exchanges) between the various
hydration shells of the polar head group
into the hydrocarbon core (Hauser and
Phillips, 1979), driven by the transmembrane water concentration energy gradient. Divalent cations compete with water
for interaction with the negatively charged
phospholipid groups and potentially displace the water molecules (Hauser et al.,
1976).
Permeability coefficients for water
movement through cell membranes vary
by orders of magnitude depending on the
membrane composition and physical state
(Finkelstein, 1978; Carruthers and Melchior, 1983). Fluid membranes are more
highly permeable than are those in a liquidcrystalline state. In fluid state membranes,
elevating temperatures, decreasing the
chain length of hydrocarbon tails of phospholipids, increasing hydrocarbon saturation, and reducing amounts of cholesterol
(within certain limits) in lipid bilayers all
act to increase water permeability. Considering the effects and molecular dimensions
of these changes, modifications of gaseous
and ionic permeability might also be
expected, albeit in complex fashion.
Whereas the lipid bilayer membrane may
represent the principal pathway of water
movement, electrolytes are virtually
impermeant. Ions and hydrophilic nonelectrolytes require a hydrophilic environ-
949
ment for movement through lipid bilayers.
Thus, permeation of cells by ions requires
integral membrane proteins in the form of
channels or carriers. The penetration of
channels by ions is dependent on such factors as the steric hindrance of molecules at
the channel entrance, the viscous drag and
charge interaction between molecules and
the walls of the channel, and the configuration of the channel length. Clearly, the
importance of these factors will depend on
the relationship between the size of the
transported molecule and the radius of the
conducting channel. Transport of ions
and water may be single-file in narrow
channels (Finkelstein, 1984). In this situation, the rate of movement of an ion such
as Na+ may be equal to that of a water
molecule moving the length of the channel. The equality of the two rates means
that the movement of water can be a major
barrier to ion transport. The exit step is
an additional barrier to the ion so that the
rate of movement of ions across the entire
membrane (channel) may be less than that
for water. Moreover, these considerations
raise the possibility that water permeability
of a channel could be salt-dependent, so
that in essence the ion can conceivably block
a channel to flow of water (Finkelstein,
1984).
The permeation of membranes by ions
is usually measured as isotopic flux or electrical conductance. Extensive literature on
this subject will not be reviewed here. Studies of integument {e.g., frog skin) indicate
that ion movement may be active or passive
and that ion transport therefore may influence the permeation of water and dissolved
gases. On the other hand, solvent drag of
electrolytes may accompany flow of water
within aqueous membrane channels.
The diffusion of gases through membranes is described by the application of
Fick's laws for diffusion and Henry's laws
for partial pressure relations and gas solubility (discussed in Feder and Burggren,
1985). Because of its high solubility, carbon dioxide is far more permeable in most
tissues than is oxygen, which accounts for
the consistent observation that cutaneous
fractional CO2 loss exceeds cutaneous oxygen uptake in bimodal, skin breathing vertebrates. Presumably, both gases permeate
950
H . B. LlLLYWHITE AND P. F. A. MADERSON
tissues wherever water movement occurs
in the bulk phase, i.e., through channels or
intercellular spaces. Additionally, respiratory gases dissolve in thin lipid membranes
at membrane-water interfaces in proportion to the partial pressures in the aqueous
phase. It is assumed that for oxygen the
boundary processes are rapid, so that the
rate limiting step is the diffusion of gas
within the membrane. Diffusion of oxygen
in the aqueous phase may, in certain circumstances, be facilitated by virtue of
microturbulence induced by the pumping
actions of respiring mitochondria or the
presence of haem or haem-like carriers (e.g.,
Lee and Smith, 1965). These factors may
account for diffusion coefficients that are
higher than that through free water
(MacDougall and McCabe, 1967).
The movement of CO2 across an epithelium involves diffusion of the dissolved CO2
gas across lipid bilayers and also reactions
among the several chemical forms of CO2
in the aqueous layers. At least two forms
of CO 2 , HCCV and CO S 2 ", cannot easily
diffuse across most cell membranes.
Numerous studies indicate that different
steps in the movement of CO2 may be rate
limiting under different conditions. In the
absence of carbonic anhydrase, the diffusion of CO2 through aqueous compartments may be rate-limiting because the
uncatalyzed hydration-dehydration of CO2
is too slow for [HCOS~] to facilitate CO2
diffusion through the aqueous layer. However, presence of carbonic anhydrase stimulates CO 2 flux 10-100-fold in proportion
to the [HCO3~] over the pH range 7-8
(Gutknecht et al., 1977). Carbonic anhydrase is widely distributed in respiratory
epithelia including the integument of a
variety of taxa (e.g., Simkiss and Wilbur,
1977; Hackman, 1984). Thus, in certain
circumstances the diffusion of CO2 through
membranes becomes the rate-limiting step.
Permeabilities of bilayer membranes to
CO2 also vary at least two orders of magnitude depending on their lipid fluidity and
composition.
CELLULAR JUNCTIONS AND THE
PARACELLULAR DIFFUSION PATHWAY
The basic organization of any epidermis
results in two pathways for transepidermal
molecular movement. The first pathway is
located between the cells (lateral, intercellular spaces), where solutes and fluids can
flow down their respective chemical and,
in some cases, electrical gradients. The second, parallel pathway is through the cells,
where movement may be by diffusion
through the lipid bilayer or by bulk flow
through channels in the cell membrane and
is potentially a multi-step process. Cellular
physiologists often categorize simple epithelia according to the relative permeability of these two pathways. In so-called tight
epithelia, junctional contacts between cells
have high resistance, so that most passive
diffusion occurs through the cell membranes. In contrast, leaky epithelia are those
in. which most diffusion occurs between
cells, that is, it is paracellular. While these
distinctions are potentially applicable to
complex, multi-layered integuments,
information is presently insufficient to
attempt such categorization.
The junctional contacts between cells are
structurally modified so that cells can
adhere, interact and dissipate tensional
stresses throughout a tissue. Additionally,
specialized junctions confer an occluding
function that allows concentration gradients to be maintained across the epidermal
layers. The term "tight junction" has been
loosely applied to a broad range of intimate
contacts between plasma membranes,
although the term was originally introduced to designate the zonula occludens
(Farquhar and Palade, 1963). The term will
here be used to designate any belt-like
region of membrane apposition which
occludes the intercellular space. Typical
tight junctions join cells at their apical edges
by a continuous belt, thus forming a planar
array.
Freeze fracture studies of tight junctions
show the presence of a network of fibrils
of diverse complexity within the membranes, thought to be the sealing elements
of the junction. In some tissues the number
of intramembrane fibrils and junctional
permeability are directly related (Claude
and Goodenough, 1973), although other
features appear to be related to the permeability properties of the junction. Often
the fibrils are not continuous but are interrupted at more or less regular intervals,
SKIN STRUCTURE AND PERMEABILITY
which might constitute leak pathways
through the junction. Differences of permeability might also depend on the chemical composition of the molecules within
the junctions rather than, or in addition
to, their morphological features. Studies
on a variety of "leaky" epithelia of endothelial origin indicate that junctional
permeabilities are larger to cations than to
anions; i.e., the tight junction constitutes a
negative environment for ion diffusion.
The relative cation- or anion-selectivity may
also change, depending on the pH of the
junction environment. Such ionic selectivity is not well characterized for tight epithelia, mainly because of the small area
occupied by the junctional pathway.
One of the principal questions relating
to permeability and transport in skin is:
What fraction of the transepithelial diffusion of substances goes through the junctional or paracellular pathway? So far, the
most reliable information on this question
comes from studies of epithelial slices of
frog epidermis which do not constitute a
normal multilayered integument. Clearly,
however, frog skin is very "tight," having
resistances of 10-50,000 ohm-cm2 compared with resistances ranging from less
than 10 to a few hundred ohm • cm2 in leakier epithelia of endothelial origin. Various
data indicate that more than 90% of the
transepithelial conductance is localized in
the paracellular pathways of leaky epithelia. By contrast, less than 10% of the total
conductance resides in the paracellular
route of tighter epithelia such as frog skin
(Erlij and Martinez-Palomo, 1978), but such
determinations are imprecise. It must be
remembered that, with respect to different
ions, epithelia are selectively permeable and
that both passive and active permeation
may result in net fluxes that are variably
partitioned between cellular and paracellular routes.
Tight junctions restrict the diffusion of
both fluids and solutes between cells. In
addition to electrolytes, various nonelectrolytes (and undoubtedly gases) are known
to permeate tight junctions, presumably
through hydrated pores. Consensus is presently lacking, however, regarding the evaluation of relative hydraulic conductivities
of the transcellular and paracellular path-
951
ways. Based on geometrical considerations,
several investigators have rejected the
notion that tight junctions are important
routes for water permeation, and some
believe that the hydraulic conductivity of
the junctional pathway can account for only
10% of the whole epithelium (Wrights a/.,
1972). However, the junctional pathway
may account for most of the water movement in "leaky" epithelia (Levitt, 1981).
Further research is required to partition
the transport of water with respect to cellular and extracellular routes, especially in
relation to varying permeability requirements of animal integuments.
In frog skin the extracellular spaces are
sealed by tight junctions in at least two layers of cells: the superficial, cornified cell
layer and the outermost cell layer of the
stratum granulosum. Recent evidence indicates that the cornified cells are ineffective
diffusion barriers and, therefore, the barrier function of cellular junctions is localized to the outside border of the outermost
cells in the stratum granulosum (Erlij and
Ussing, 1978). Undoubtedly, however, the
permeability properties of the cellular
junctions are, in part, functions of the
osmotic state of the cells, which depends
on the permeability of the outermost skin
and its mucous covering (see below).
Changes of cell volume induce changes in
active transport (Erlij and Ussing, 1978) in
addition to altering the structure of intercellular spaces. Changes in the morphology of intercellular spaces can affect
diffusion of water and solutes even independently of cellular swelling or shrinkage.
For example, osmotic flow of water within
epithelia can cause dilation or collapse of
lateral extracellular spaces (depending on
the flow direction). Calculations show that
diffusion through these spaces can become
rate-limiting as they collapse and that such
changes in the dimensions of lateral spaces
can produce asymmetry of water flow
(Wright*/ al., 1972).
Other factors can modify the properties
of the paracellular pathway. Low pH, for
example, weakens intercellular junctions
thus leading to increased paracellular permeation by ions (Ferreira and Hill, 1982;
Marshall, 1985).
Most investigations of cellular junctions
952
H . B. LlLLYWHITE AND P. F . A. MADERSON
have been restricted to vertebrate tissues
and to invertebrate tissues of endothelial
origin. Tight junctions are absent from the
vast majority of invertebrate tissues, being
replaced by septate junctions. These are
structures that, in transverse sectional view,
comprise a series of septa spanning 15-18
nm intercellular spaces. Septate junctions
occur in all invertebrate integuments studied to date and always form a belt around
the apical edges of cells lining the outer
epithelia. Circumstantial evidence suggests
that they restrict paracellular diffusion in
a manner that is functionally analogous to
that of vertebrate tight junctions (Green,
1984).
Circumapical tight junctions are present
in all classes of vertebrates, although they
appear to occur sparsely in skin of mammals (Matoltsy, 1984). It is not clear
whether the emphasis placed on tight junctions in amphibian skin merely reflects the
extensive use of that system as a model for
study of membrane permeability.
STRUCTURAL AND SECRETED BARRIERS
The epidermis of numerous invertebrates with an exposed plasma membrane
surface is primarily an absorptive, transporting organ. Primitively, as well as in specialized forms such as parasitic tapeworms,
the integument is important for nutrient
absorption in addition to ion regulation or
gaseous exchange. During the evolutionary radiation of eumetazoan animals, a
variety of factors (including increasing size,
terrestriality and mobility) necessitated
structural reinforcement of the skin for
protection and support. In general terms,
this evolution proceeded from a flagellated
or ciliated epidermis to a "hypodermis"
covered with a secreted cuticle. Although
primitive cuticles may have regulated the
net flux of nutrients as a primary function,
their appearance and radiation was often
correlated with skeletal, protective or locomotory needs of the organism (Rieger,
1984).
The differentiation of a true cuticle,
completely absent from vertebrates,
involves the secretion of fibrous and/or
granular materials onto the outer surface
of the epidermis. However, supportive
fibrous and granular materials may also be
associated with the base of the epidermal
cells or with the apical cytoplasm. In more
advanced eumetazoans, the fibrous matrix
of the cuticle may become specialized by
division into distinct layers in addition to
the incorporation of chitin, collagen and
minerals. Various acids, mucopolysaccharides and lipids may also form associations
with the fibrous matrix.
Chitin is a high molecular mass polysaccharide that forms a fibrous component of
many cuticles and confers tensile strength
combined with flexibility. The chitin molecule is long and straight and is stabilized
by a covalently bonded backbone and
intramolecular hydrogen bonds between
adjacent residues along the chain. Microfibrils of chitin associate with many different proteins having a wide range of molecular mass. Interface bonding between chitin
and the immediately neighboring protein
is strong. The chitin fibers of insect cuticle
lie on a matrix of cross-linked protein,
forming a system which has a large strength
to mass ratio and high tensile and compressive strengths that are stronger than
either of the two components alone. Several crystalline forms of chitin are known,
commonly forming lattices that are
arranged in the manner of a plywood. The
ratio of protein to chitin varies widely, but
together these two components normally
account for more than 90% of the organic
content of cuticle. In sclerotized cuticles
mineralization involves calcium or magnesium carbonates and phosphates whose
quantity is inversely related to the protein
content. Mineral deposition occurs both
within and between fibrils and may be crystalline in form.
Invertebrate collagens and vertebrate
keratins also have two-phase structures with
helicoidal arrangements present in some of
the fiber layers. The strength of these systems is increased by having fibers of high
tensile strength and elasticity oriented in a
matrix of weaker proteins or polysaccharides. Keratin complexes are heterogeneous collections of" proteins containing
many sulphydryl (SH) and disulphide (SS)
linkages of cystine. These form the main
constituents of the dead horny cells in
SKIN STRUCTURE AND PERMEABILITY
superficial layers of epidermis and its derivatives (hair and feathers) in amniote vertebrates. They may be associated with
fibrous or globular proteins, variations that
allow wide differences in structure and
mechanical properties of keratinized cells.
Mineralization of horny tissue involves calcium that is usually bonded to phospholipid within the keratin complex.
Both fibrous cuticle and subepidermal
tissue must be traversed by diffusion, so
these structures may represent a significant
resistance to mass transfer, as compared
with the epidermal resistance. Keratins of
vertebrates and cuticles of invertebrates
vary from thin, transparent membranes to
thick, tough and rigid armour. In very general terms, permeabilities of fibrous structures are related to their thickness, and
where hard and soft tissues occur on the
same body surface, the important sites of
exchange are at the soft tissue. Thick, calcified or sclerotized structures such as the
shells of molluscs, exoskeleton of crustaceans and calcareous ossicles of echinoderms are virtually impermeable to gases,
ions and water. At the other extreme, thin
and highly permeable cuticles may cover
exchange structures such as gills or occur
on the general body surface immediately
after molting (e.g., Mangum et al., 1985).
Thickness is not an absolute predictor of
permeability, however, because the dimension is not independent of the tissue composition (see Lillywhite and Maderson,
1982).
How do the structural features affect the
resistance of fibrous polymers to molecular
diffusion? From a strictly mechanistic viewpoint, any alteration in a polymer structure
that affects the free volume (equivalent to
"holes" or "pores") should alter permeability to invading molecules. Moreover,
permeating molecules will have permeability coefficients related to their size,
shape and polarity. Due to steric hindrance, the diffusion coefficient is expected
to decrease considerably with increasing
molecular mass of the diffusing substance.
Studies of avian eggshells demonstrate that
their permeability to O 2 , CO2 and water
vapor varies in direct proportion to the
differences in diffusion coefficients, sug-
953
gesting a common diffusion pathway for
gaseous molecules (Paganelli et al., 1978;
Ackerman and Rahn, 1980).
Generally, studies on permeability of
polymers, including collagen, show that
alterations of side chains, crystallinity, polar
groups, plasticizers and fillers all affect permeability (Lieberman et al, 1972). With
respect to side chains, permeability is
affected below a critical molecular mass of
fiber cross-linkages. Additionally, crosslinking beyond certain critical levels
increases the resistance to diffusion. Stability of the cross-linkage structure is
dependent on a large number of intermolecular forces: mobility and free volumes of biological polymers are significantly affected by covalent, ionic and
hydrogen bonding as well as van der Waals
attractive forces between nonpolar amino
acid side chains.
Proteins that are sclerotized are hardened and stabilized by the presence of aromatic cross-links, which decrease permeability. Hardening is also attributable to
mineralization in some species. Many crustacean cuticles are both calcified and sclerotized. Proteins from nonsclerotized cuticles generally have a higher content of
amino acids with bulky side chains and so
inhibit close packing of the molecules. The
woodlouse cuticle, for example, is relatively permeable compared with that of
insects. X-ray diffraction data suggest that
the proteins may be globular with a high
proline content, which limits the degree of
helix formation and packing that is possible (reviewed in Hackman, 1984). In contrast, proteins from sclerotized cuticles have
generally smaller and nonpolar amino acids
that are able to pack more closely together,
thereby permitting formation of many van
der Waals bonds.
Hydration significantly affects the permeability of fibrous protein polymers
because of the influence of water on the
molecular structure. Proteins bind water
very strongly at low relative humidities, and
the water content increases in proportion
to humidity exposure. The mobility of side
chains and the distance between them
increases with increasing water content as
water enters between the protein mole-
954
H . B. LlLLYWHITE AND P . F. A. MADERSON
cules and reduces the degree of crystallinity. Introduction of water molecules into
the polymer structure increases the available free volume as well as the mobility of
cross-linkage groups. Dehydration reduces
permeability not only by reversing these
effects but also by promoting noncovalent
bondings between protein chains (Vincent
and Hillerton, 1979). Consequently, the
permeability of dry protein films is
extremely low (Lieberman et al., 1972).
Obviously the pH of fibrous structures
is significant with respect to ion diffusion,
the stability of molecular associations, and
the bonding of water. Arthropod cuticles
tend to be maintained at a pH close to the
isoelectric points of their proteins, such that
cross-linkages between protein chains are
strongest. Marked deviations of pH from
these isoelectric points potentially alter the
water content of cuticles because bondings
between chains are replaced by bondings
between protein and water (Hackman,
1984). Ion exchange can be markedly
affected in some cuticles which have a
matrix including acid polysaccharides
(Gomme, 1984).
Fluid secretions may contribute to the
cuticle of various animals, especially in taxa
lacking a fibrous or mineralized covering
of significant thickness. These secretions
serve a variety of functions including
mechanical protection, pH regulation, ion
exchange, antibiotic actions, friction
reduction, pheromonal roles and prevention of dehydration. Most secretions are
generally mucous or proteinaceous in varying proportions. Mucous secretions are
predominantly water with varying amounts
of proteoglycans and glycoproteins (glycoconjugates), various ions and often lesser
quantities of sugars, amino acids and lipids
(Campbell et al., 1967; Wilson, 1968; Dapson, 1970). Secretions may be either acidic
or alkaline, the latter possibly counteracting acidification of the skin due to CO2
excretion (Friedman et al., 1967).
In fundamental terms, an epicutaneous
film of mucus may be regarded as an
unstirred layer whose resistance to diffusion is proportional to the thickness, viscosity and solute concentration. Mucus
covering the skin of carp, Cyprinus carpio,
has an oxygen diffusivity that is about 70%
that of the surrounding water (Ultsch and
Gros, 1979). However, very thin films are
typically involved where the underlying tissue is absorptive or respiratory, and a variety of evidence indicates that such films do
not limit diffusion significantly. The secretions themselves are sources of water, solute and gaseous efflux, although water and
electrolytes might be recycled at the skin
surface (Machin, 1977; Whitear, 1977).
Mucous secretions are important in preventing dehydration of exposed epidermal
surfaces in dehydrating environments suggesting a fundamental dichotomy of cuticular organization in terrestrial animals.
With few exceptions, either the epidermis
is provided with a lipid and/or fibrous diffusion barrier to prevent excessive water
loss from the skin, or the epidermis is covered with a wet film so that evaporation
occurs from secreted fluids rather than
from the epidermal cells. In terrestrial gastropods and amphibians, mucus secretion
is a fundamental mechanism for the transfer of water to evaporating surfaces, which
are sculptured to spread and retain the fluid
(Machin, 1964a, b; Lillywhite, 1971). Without mucus extrusion, the diffusional permeability of the epidermis is unable to sustain normal levels of evaporative water loss,
and the skin dries (see also Lillywhite,
1975). With respect to cutaneous gas
exchange, it is not necessary for oxygen or
CO2 to dissolve in water before entering
or leaving the outer epidermal membrane
if the latter is exposed to air. Rather, the
maintenance of "wet" respiratory structures is essential to preserve the functional
morphology of the diffusion membrane,
including patency of its perfusion vessels.
This point is often not clearly represented
in textbooks.
LIPIDS AND WATER DIFFUSION:
BIOCHEMICAL BARRIERS
Surveys indicate that numerous terrestrial arthropods and vertebrates have lost
the ability to exchange gas and solutes
through the integument by developing a
protective and relatively waterproof epidermis. The maintenance of internal water
volume and concentration is an important
SKIN STRUCTURE AND PERMEABILITY
problem faced by all animals, but one that
is magnified in terrestrial species, which
may have limited access to water and live
in dehydrating conditions. Mechanisms for
reducing the diffusion of water through
skin are varied and not entirely understood; however, the most effective reduction of water permeability is achieved in
those integuments where lipids are deposited as a more or less continuous sheet. It
appears that the "tight" nature of such a
diffusion barrier eminently affects gases and
solutes as well as water.
Lipoid materials occur at various places
throughout all integuments. However, the
use of lipids to form layered, watertight
barriers is best known in certain arthropods and vertebrates where the resistance
mechanisms have been well studied (see
Hadley, 1980, 1981; Lillywhite and Maderson, 1982 for reviews).
Evidence suggests that diverse chemical
compounds in all of the cuticular layers
may contribute to the impermeability of
integument in terrestrial arthropods. But
the principal recognized water barrier in
most insects and arachnids consists of lipid
layers or impregnated waxes associated
with the epicuticle. In particular, the
importance of surface lipids in restricting
cuticular water loss is firmly established.
Water efflux from the cuticle increases significantly when these lipids are removed
by solvent extraction or mechanical abrasion. Many studies have shown correlations
between the quantity of extracted cuticular
lipids, which may vary seasonally as well as
interspecifically, and their effectiveness as
a diffusion barrier. This relationship has
exceptions however, and compositional
features of lipids are also important.
Chemical analyses indicate that cuticular
lipids of most insects and arachnids consist
of a heterogeneous mixture of compounds
with long chain hydrocarbons predominating (Hadley, 1981). Free fatty acids, wax
esters, cholesterol and other categories of
lipid are usually present in smaller amounts,
whereas alcohols and phospholipids are
often not present. Lipid barriers exhibiting the greatest resistance to water flux are
characterized by predominance of largely
nonpolar, long-chain, saturated and usu-
955
ally branched alkanes that appear not to
be organized into specifically oriented layers (see also Hadley, 1984). The long chain
molecules pack closely and increase the
intensity of van der Waals interactions
between hydrocarbon molecules, thus creating a nonfluid membrane that resists
penetration by water.
Lipids are present in the epidermis of all
classes of vertebrates, although their influence on permeability is not significant in
most amphibians and is not well studied in
birds. Some species of South American
phyllomedusine frogs secrete lipids onto
the skin surface and then use their limbs
to spread the secretion over the body (Blaylock et al., 1976). The composition of these
lipids has been analyzed for one species,
Phyllomedusa sauvagei, and was found to
contain largely long-chain wax esters
(McClanahan et al., 1978). The superficial
film of lipid retards water loss while frogs
are inactive in trees where they are exposed
to the (usually dry) atmosphere.
Numerous reptiles live exposed to dry
conditions and generally such species possess quite impermeable integuments. Variation in the integumentary water permeability of squamate reptiles is correlated
with habitat and determined more by lipid
composition of the epidermis than by the
thickness or structure of its keratin (Lillywhite and Maderson, 1982). The lipid
barrier resides principally in the mesos
layer, which is overlain by layers of beta
keratin whose role is primarily structural
(Fig. 1). The mesos lipids comprise a complex mixture of polar and neutral lipids
which appear to be similar in composition
to those comprising the diffusion barrier
of mammalian epidermis (see below). The
mesos lipids occur as intercellular sheets
and are derived from lamellar granules that
extrude their contents into the intercellular space (Landmann, 1986). Part of the
contents remain within mesos cells while
the expelled lipids envelop the cells from
all sides and fill the intercellular spaces to
form a continuous sheet.
A similar process occurs in mammalian
epidermis, wherein lipids are deposited
intercellularly from lamellar granules situated at the boundary of the stratum gran-
956
H. B. LlLLYWHITE AND P. F. A. MADERSON
ulosum and stratum corneum. The expelled
contents of the granules form broad lipid
sheets that lie parallel with the envelopes
and filaments of keratinized cells. The
major group of lipids that contribute to
these extracellular sheets are highly saturated, unbranched, long-chain ceramides
(Wertz, 1986). The high degree of saturation resists oxidative damage, and the
paucity of branching makes these molecules capable of forming tightly packed,
nearly crystalline arrays of hydrophobic
covering.
Does a lipoid water barrier necessarily
impede diffusion of other important molecules such as respiratory gases and ions?
Information is conflicting on this point, and
a satisfactory understanding of the mechanistic constraints governing independent
adjustments of permeability to water, ions
and gases must await further research that
specifically addresses these interactions.
Damage to the cuticle of insects appears to
affect water permeability more so than permeability to gases, and the waterproofing
waxes of insect eggs are reported to be very
permeable to oxygen (reviewed in Buck,
1962). Also, in aquatic reptiles permeability of skin to water and gases does not necessarily parallel permeability to electrolytes. In addition, there is an apparent
asymmetry in the diffusion of water and
electrolytes through the skin, which can be
altered by extraction of lipid (Stokes and
Dunson, 1982; Dunson and Stokes, 1983).
These findings are controversial, but at
least a circumstantial case can be made suggesting that permeabilities of skin to different substances may have some independent components of adjustment.
On the other hand, numerous surveys
indicate that highly water impermeable
integuments are also poor exchangers of
gases and ions. Moreover, strong correlations between water loss and CO2 loss during ventilatory activity of insects {e.g.,
Quinlan and Hadley, 1982) are indicative
of the dual resistance of cuticle to both CO2
and water. Recently, pulmonary and cutaneous gas exchange and cutaneous evaporative water loss were measured simultaneously in the xeric-adapted frog
Phyllomedusa sauvagei (Stinner and Shoe-
maker, unpublished data). Evaporative
water loss varied widely between individual
frogs, presumably due to variation in the
thickness or distribution of the waterproofing lipids that covered the outermost epidermis. The interesting finding in the present context was that cutaneous oxygen
uptake and CO2 loss varied directly with
the magnitude of cutaneous water loss (Fig.
2). These data indicate that reducing cutaneous water loss by use of a lipid barrier
entails an attendant reduction in cutaneous
gas exchange.
VASCULAR MORPHOLOGY AND
PERFUSION
Perfusion of the skin with blood is essential to integumentary function in all higher
animals characterized by a well-organized
circulation. Blood circulation not only services the metabolic needs of active cutaneous tissue, but also provides the transport medium for substances exchanged at
the skin surface. Morphological aspects of
integumentary perfusion are fundamental
to the regulation of internal fluid composition, as the capillary architecture determines both the functional area and the distance through which diffusion and
exchange occur.
The morphology of cutaneous vasculature has been thoroughly studied in comparatively few species. Although great
variation exists, several generalizations
concerning vascular patterns are evident.
Vascularization is essentially a property of
the dermis, and capillaries rarely penetrate
the epidermal structures. Presumably,
extensive vascularization of epidermis
compromises the barrier and protective
functions of the skin. In mammals, capillary units extend as a "candelabra" of loops
projecting vertically to contact the basal
membrane and form papillary intrusions
of the epidermis. These drain into venules
arranged horizontally in units that tend to
outnumber the arterioles. Endothelium
adjacent to the epidermis is fenestrated and
sometimes associated with microfilaments
or myofilaments.
Other vertebrates display variations of
this general scheme. Often, there is an
elaborate network of anastomosing vessels
957
SKIN STRUCTURE AND PERMEABILITY
60r
•
50 -
•
•
to
40 • •
UJ
30
0
•
I
O
x
UJ
CO
•
0,
20 ^. —
yS
10 -
O <fi
•cr
JO-
—
0
0
-tr
1
5
CUTANEOUS
10
WATER
15
LOSS
1
20
(mg-g-i-h"1)
1
25
FIG. 2. Relationship between cutaneous gas exchange and cutaneous evaporative water loss in the frog
Phyllomedusa sauvagei. Measurements were made as frogs rested with variably groomed layers of lipids that
were secreted from integumentary glands onto the skin surface. Rates of oxygen uptake (open circles) and
CO2 loss (dark circles) are indicated as percentages of the total oxygen or CO2 exchange (pulmonary plus
cutaneous flux). Data are from J. Stinner and V. Shoemaker, unpublished (with permission).
contained within loose connective tissue of
the superficial dermis which has a low
nutritional requirement. In bony fishes,
there is a secondary microvascular system
immediately below the epidermis but overlying the scales and corresponding to their
topography (Vogel, 1985). The cutaneous
vascular systems of squamate reptiles
underlie the keratinized elements of the
outer scale surfaces but are extensively
developed within the inner and outer scale
regions as well as the free edges of scales
(Drane and Webb, 1980). The various patterns that are seen very often suggest that
the cutaneous vascular system is functionally related to exchange processes between
the animal and its environment. Capillary
networks are often less developed in regions
of skin that are less exposed to environmental medium or are subjected to excessive abrasion.
Among the more extreme specializations of cutaneous vasculature are those
that have developed in relation to respiratory and nutritional functions in invertebrates. For example, the tentacles of
pogonophores bear pinnules consisting of
single epidermal cells that are elongated
distally. The cuticle covering the pinnule
is nonfibrous and exceedingly thin, while
the inner cytoplasm is largely occluded by
two blood capillaries which unite near the
tip of the cell. Outgrowths of these capillaries associate with cisternae of the smooth
endoplasmic reticulum in some species. The
capillaries are essentially extracellular,
however, inasmuch as they are formed by
invaginations of the basal plasma membrane and basal lamina which connect with
blood spaces between the epidermal cells
and subjacent muscle cells (George, 1977).
Some of the cutaneous blood vessels of
earthworms are similarly lined with extracellular basement membrane (Hama,
1969). These vascular specializations
doubtless have a respiratory role.
The pogonophore example involves both
morphological reduction of the cuticle and
modification of the blood transport system
as means of reducing the diffusion path
between the environment and internal fluid
spaces. Epithelial thinning is also apparent
in various vertebrates in which cutaneous
gas exchange is demonstrated to be impor-
958
H . B. LlLLYWHITE AND P. F. A. MADERSON
tant. As noted by others (e.g., Feder and
Burggren, 1985), the diffusion barrier of
specialized epithelia in fish gills may be
fractions of a ^m, whereas diffusion barriers of the general integument are greater,
reflecting the protective and supportive
functions that evidently must be accommodated by the cutaneous morphology. In
many amphibians the epidermal diffusion
barrier separating blood from the outside
medium varies from about 12 /im in lungless salamanders to more than 60 /wn in
some bufonids, which, is adequate to support diffusional exchange (Czopek, 1965).
Cutaneous capillaries may be deeper in the
skin of xeric frogs (Drewes et al., 1977),
whereas in frogs (Telmatobius) or salamanders (Cryptobranchus) that rely on cutaneous gas exchange the skin vasculature is
very superficial and may penetrate the epidermis (Rabl, 1931; Guimond and Hutchison, 1973; Hutchison et al., 1976).
Epidermal thickness in the skin of fishes
varies considerably, even among species
that rely on cutaneous gas exchange (Mittal
and Munshi, 1971). Both eels and pond
loaches can supply nearly the whole of their
oxygen demand cutaneously, although the
epidermis is 263 fim thick in Anguilla
anguilla (Jakubowski, 1960) and 339 nm
thick in Misgurnus fossilis (Jakubowski,
1958). On the other hand, the epidermis
of the air-breathing fish Mastacembelus pancalus may be as thin as 7 jtm at places where
blood capillaries penetrate the epidermis
(Mittal and Munshi, 1971). This latter
species occupies stagnant pools that are
subject to drying, and the cutaneous morphology is presumably an adaptation that
facilitates cutaneous oxygen extraction
when the fish is exposed to air.
Both density of capillaries and the diffusion distance (epidermal thickness) that
overlies them show variability related to
demands for exchange processes. Amphibians and fishes that inhabit moist or aquatic
environments and are variously dependent
on cutaneous transfer of respiratory gases
possess dense networks of skin capillaries
(reviewed in Randall, 1970; Feder and
Burggren, 1985). However, there is no
consistent relationship between the extent
of cutaneous vascularization, thickness of
the epidermis, and the amount of oxygen
uptake occurring across the skin. This is
because the exchange process must take
into account the total functional surface
area of skin with capillaries, the diffusion
gradient, blood flow and oxygen loading
characteristics, metabolic requirement for
oxygen, and competing requirements that
also shape the skin morphology. Moreover,
integumentary structure and vascularization may vary regionally and through time.
Both theoretical and empirical studies
have shown that molecular flux of a substance having a low permeability coefficient is independent of blood flow and thus
governed largely or exclusively by the concentration gradient and permeability constant. However, as permeability increases,
movement or uptake becomes more
dependent on blood flow. Therefore, considering the movement of a molecular
species across the skin, two factors in combination limit the transport between blood
and the external environment: (a) diffusion
limitation, which is the resistance of the
epithelium to molecular transport, and (b)
perfusion limitation, which is the lack of
molecules available for transport across or
away from the epithelium as a result of
insufficient blood flow. A limited but growing number of studies have provided indices
of diffusion limitation and perfusion limitation, largely with respect to the mass
transfer of water and respiratory gases in
amphibian integument and noncutaneous
epithelia.
Studies of skin from Bufo bufo and Rana
pipiens indicate that both osmotic and diffusive water exchange are perfusiondependent when skin permeability is high
(Christensen, 1974, 1975; Mahany and
Parsons, 1978). Thus, when skin permeability is increased with arginine vasotocin
(ADH), perfusion rate markedly affects
water flux through the cutaneous tissue.
Moreover, while arginine vasotocin
increases osmotic water exchange substantially, diffusive water transfer is always more
dependent on blood flow than on ADHinduced permeability changes. Similarly,
water exchange is shown to be dependent
on blood flow in other epithelia such as gut
and muscle (e.g., Dobson et al., 1971).
SKIN STRUCTURE AND PERMEABILITY
Studies of cutaneous gas exchange in a
variety of amphibians have demonstrated
diffusion limitations in addition to perfusion-dependent enhancement of the skin
diffusing capacity. Inasmuch as these findings are reviewed elsewhere, extensive
comments will not be made here (see Feder
and Burggren, 1985; Malvin, 1988; Piiper,
1988). As in studies of water permeability,
the regulation of cutaneous gas exchange
involves a complex interaction of multiple
factors involving both diffusion and perfusion limitations. Perfusion dependent
changes of the skin diffusing capacity are
often related to conditions in the ambient
medium, including the ventilatory disturbance of external unstirred layers (Feder
and Pinder, 1988). However, a comprehensive picture of how perfusion is regulated in relation to diffusion limitations
must remain a goal for future research,
especially in contexts of conflicting permeability requirements for different permeating substances.
Clearly, blood flow can alter the mass
transfer of substances in several ways. (1)
Increasing the rate of flow in patent, perfused capillaries potentially increases
transfer but may have little or no effect in
diffusion limited systems (Piiper, 1988). (2)
If the increment of perfusion involves
increasing the number of perfused capillaries (recruitment), then the functional
exchange surface is increased. Capillary
recruitment has been shown to enhance
gas transfer across integument of amphibians and humans (Feder and Burggren,
1985). (3) Considering the morphology of
cutaneous vasculature, capillary recruitment might also entail reductions in diffusing distance as well as increases in diffusing area. The relative contributions of
these two aspects remain to be quantified,
however. (4) It is conceivable that increases
in perfusion can modify mass transfer rates
indirectly, and this might apply in several
exemplary situations. In the case of ion
transport, one explanation for dependence
of sodium flux on perfusion flow rate is
that sodium is actively transported and
some component of this movement is sensitive to perfusate oxygen or metabolite
availability (Claiborne and Evans, 1981).
959
As a related example, blood flow changes
may indirectly affect cutaneous oxygen
uptake through an effect on metabolism of
skin tissue. In some species of fish, cutaneous oxygen consumption equals or
exceeds the cutaneous oxygen uptake from
the external medium (Kirsch and Nonnotte, 1977). Consequently, the diffusion
gradient for oxygen across the epidermis
may be greatly influenced by the level of
mitochondrial respiration in the skin. (5)
In terrestrial species, blood flow conceivably affects the hydration status of fibrous
proteins and thus the free space available
for molecular diffusion within the fibrous
matrix (see p. 953). (6) Finally, changes of
perfusion patterns are potentially important with respect to the timing and geometry of skin ventilation. Studies have shown
that larvae of air-breathing teleosts have
cutaneous blood flow that runs countercurrent to the water stream (Liem, 1981).
Thus, regulation of skin perfusion might
be important with respect to the timing
and frequency of movements of the pectoral fins that generate posteriorly directed
respiratory water flow. Conceivably, regulation of cutaneous ventilation and perfusion matching, if only at a more localized
scale, might be part of a complex pattern
of responses in other species of animals.
CONCLUSION
We have discussed the structural attributes of integument as they relate to
exchange processes involving gases, ions
and water. Unlike simple membranes on
which the understanding of many mechanistic principles is based, animal integuments are heterogeneous, multimembrane
systems capable of regulating exchange
pathways and barriers for numerous substances in relation to widely different and
sometimes changing environmental
demands. It is possible to quantify the
transport or diffusing capacity of a given
integument for a particular substance, using
a "black box" approach. However, as noted
byjorgen Gomme (1984), "A comprehensive understanding of the integument as a
barrier to and pathway of communication
with the environment can be achieved only
when a comparative study of the individual
960
H . B . LlLLYWHITE AND P. F. A. MADERSON
transfer processes is integrated into an
analysis of their individual regulation and
mutual interdependency." Evolutionary
considerations and broad surveys of the
occurrence and function of junctional
complexes and fibrous, mineralized or
lipoid barriers emphasize the generality of
diffusion limitations. On the other hand,
insight into the characteristics of individual permeation processes suggest some
scope for independent adjustment of permeability for specific substances. The challenge for contemporary and future
research is for comparative investigations
in functional cutaneous morphology to elucidate meaningful understanding of diffusional pathways, the rate limitations for
specific molecules, and the variation that
is possible both physiologically and through
evolutionary time.
ACKNOWLEDGMENTS
We express our thanks to Martin Feder
and David Evans for helpful comments
during the preparation of the manuscript.
H.B.L. was supported by the National
Institutes of Health research grant number
HL33821 while this work was prepared.
P.F.A.M. was supported partly from PSCCUNY funds.
REFERENCES
Ackerman, R. A. and H. Rahn. 1980. In vivo O 2 and
water vapor permeability of the hen's eggshell
during early development. Respir. Physiol. 45:
1-8.
Bereiter-Hahn, J., A. G. Matoltsy, and K. S. Richards,
(eds.) 1984. Biology of the integument. 1. Inverte-
brates. Springer-Verlag, New York.
Bereiter-Hahn, J., A. G. Matoltsy, and K. S. Richards,
(eds.) 1986. Biologyojthe integument. 2. Vertebrates.
Springer-Verlag, New York.
Blaylock, L. A., R. Ruibal, and K. Platt-Aloia. 1976.
Skin structure and wiping behavior of phyllomedusine frogs. Copeia 1976:283-295.
Buck, J. 1962. Some physical aspects of insect respiration. Ann. Rev. Entomol. 7:27-56.
Campbell, J. P., R. M. Aiyawar, E. R. Berry, and E.
G. Huf. 1967. Electrolytes in frog skin secretions. Comp. Biochem. Physiol. 23:213-223.
Carruthers, A. and D. L. Melchior. 1983. Studies of
the relationship between water permeability and
bilayer physical state. Biochemistry 22:57975807.
Cass, A. 1968. Water and ion permeability of thin
lipid membranes. Ph.D. Diss., Rockefeller University, Xew York.
Christensen, C. V. 1974. Effect of arterial perfusion
on net water flux and active sodium transport
across the isolated skin of Bufo bufo bufo (L.). J.
Comp. Physiol. 93:93-104.
Christensen, C. V. 1975. Effects of dehydration,
vasotocin and hypertonicity on net water flux
through the isolated, perfused pelvic skin of Bufo
bufo bufo (L.). Comp. Biochem. Physiol. 51A:710.
Claiborne.J. B. and D. H. Evans. 1981. The effect
of perfusion and irrigation flow rate variations
on NaCl efflux from the isolated, perfused head
of the marine teleost, Myoxocephalus octodedmspi-
nosus. Marine Biol. Letters 2:123-130.
Claude, P. and D. A. Goodenough. 1973. Fracture
faces of zonulae occludentes from "tight" and
"leaky" epithelia. J. Cell Biol. 58:390-400.
Czopek, J. 1965. Quantitative studies on the morphology of respiratory surfaces in amphibians.
Acta Anat. 62:296-323.
Dapson, R. W. 1970. Histochemistry of mucus in the
skin of the frog, Rana pipiens. Anat. Rec. 166:
615-626.
Dobson, A., A. F. Sellers, and S. O. Thorlacius. 1971.
Limitation of diffusion by blood flow through
bovine ruminal epithelium. Amer. J. Physiol. 220:
1337-1343.
Drane, C. R. and G. J. W. Webb. 1980. Functional
morphology of the dermal vascular system of the
Australian lizard Tiliqua scincoides. Herpetologica
36:60-66.
Drewes, R. C , S. S. Hillman, R. W. Putnam, and O.
M.Sokol. 1977. Water, nitrogen and ion balance
in the African treefrog Chiromantis pelersi Boulenger (Anura: Rhacophoridae) with comments
on the structure of the integument. J. Comp.
Physiol. 116:257-268.
Dunson, W. A. and G. A. Stokes. 1983. Asymmetrical diffusion: A reversal of sodium and water
movement through the skin of sea snakes. Physiol. Zool. 56:106-111.
Erlij, D.and A. Martinez-Palomo. 1978. Role of tight
junctions in epithelial function. In G. Giebisch,
D. C. Tosteson, and H. H. Ussing (eds.), Membrane transport in biology. III. Transport across multi-
membrane systems, pp. 27-53. Springer-Verlag,
New York.
Erlij, D. and H. H. Ussing. 1978. Transport across
amphibian skin. In G. Giebisch, D. C. Tosteson,
and H. H. Ussing (eds.), Membrane transport in
biology. III. Transport across multi-membrane systems,
pp. 175-208. Springer-Verlag, New York.
Farquhar, M. G. and G. E. Palade. 1963. Junctional
complexes in various epithelia. J. Cell Biol. 17:
375-412.
Feder, M. E. and W. W. Burggren. 1985. Cutaneous
gas exchange in vertebrates: Design, patterns,
control and implications. Biol. Rev. 60:1—45.
Feder, M. E. and A. W. Pinder. 1988. Ventilation
and its effect on "infinite pool" exchangers. Amer.
Zool. 28:973-983.
Ferreira, K. T. G. and B. S. Hill. 1982. The effect
of low external pH on properties of the paracellular pathway and junctional structure in isolated
frog skin. J. Physio!. 332:59-67.
SKIN STRUCTURE AND PERMEABILITY
961
Finkelstein, A. 1978. Lipid bilayer membranes: Their
mined by D2O and H 2 O. Scand. Arch. Physiol.
permeability properties as related to those of cell
72:199-214.
membranes. In T. E. Andreoli, J. F. Hoffman,
Hutchison, V. H., H. B. Haines, and G. Engbretson.
and D. D. Fanestil (eds.), Physiology of membrane
1976. Aquatic life at high altitude: Respiratory
disorders, pp. 205-216. Plenum Medical Book Co.,
adaptation in the Lake Titicaca frog, Telmatobius
New York.
culeus. Respir. Physiol. 27:115-129.
Finkelstein, A. 1984. Water movement through Jakubowski, M. 1958. The structure and vascularimembrane channels. In F. Bronner and W. D.
zation of the skin of Pond-loach (Misgurnusfossilis
Stein (eds.), Current topics in membranes and transL.). Acta Biol. Cracov. (Zool.) 1:113-127.
port, Vol. 21, pp. 295-308. Academic Press, New Jakubowski, M. 1960. The structure and vasculariYork.
zation of the skin of the eel (Anguilla anguilla L.)
Finkelstein, A. and A. Cass. 1968. Permeability and
and the viviparous blenny (Zoarces viviparus L.).
electrical properties of thin lipid membranes. J.
Acta Biol. Cracov. (Zool.) 3:1-22.
Gen. Physiol. 52:145s-l72s.
Kirsch, R. and G. Nonnotte. 1977. Cutaneous resFriedman, R. T., N. S. Laprade, R. M. Aiyawar, and
piration in three freshwater teleosts. Respir.
E. G. Huf. 1967. Chemical basis for the H+ graPhysiol. 29:339-354.
dient across frog skin. Amer.J. Physiol. 212:962- Koefoed-Johnsen, V. and H. H. Ussing. 1953. The
972.
contribution of diffusion and flow to the passage
George, J. D. 1977. The pogonophore epidermis, its
of D2O through living membranes: Effect of neustructure, functions and affinities. Symp. Zool.
rohypophysial hormone on isolated anuran skin.
Soc. Lond. 39:195-222.
Acta Physiol. Scand. 28:60-76.
Gomme.J. 1984. Permeability and epidermal trans- Krogh, A. 1919. The rate of diffusion of gases
port. In J. Bereiter-Hahn, A. G. Matoltsy, and K.
through animal tissues, with some remarks on the
S. Richards (eds.), Biology of the integument. 1.
coefficient of invasion. J. Physiol. 52:391-408.
Invertebrates, pp. 323-367. Springer-Verlag, New Landmann, L. 1986. Epidermis and dermis. In J.
York.
Bereiter-Hahn, A. G. Matoltsy, and K. S. RichGreen, C. R. 1984. Intercellular junctions. In J. Beards (eds.), Biology of the integument. 2. Vertebrates,
reiter-Hahn, A. G. Matoltsy, and K. S. Richards
pp. 150-187. Springer-Verlag, New York.
(eds.), Biology of the integument. 1. Invertebrates, pp. Lee, D. L. and M. H. Smith. 1965. Hemoglobins of
5-16. Springer-Verlag, New York.
parasitic animals. Exp. Parasit. 16:392—424.
Guimond, R.W. and V.H.Hutchison. 1973. Aquatic Levitt, D. G. 1981. Routes of membrane water transrespiration: An unusual strategy in the Hellbenport: Comparative physiology. In H. H. Ussing,
der Cryptobranchus alleganiensis alleganiensis (DauN. Bindslev, N. A. Lassen, and O. Sten-Knudsen
din). Science 182:1263-1265.
(eds.), Water transport across epithelia, Alfred BenGutknechtJ., M. A. Bisson, and F. C. Tosteson. 1977.
zon Symposium 15, pp. 248-257. Munksgaard,
Diffusion of carbon dioxide through lipid bilayer
Copenhagen.
membranes: Effects of carbonic anhydrase, bicarbonate, and unstirred layers. J. Gen. Physiol. 69: Lieberman, E. R., S. G. Gilbert, and V. Srinivasa.
1972. The use of gas permeability as a molecular
779-794.
probe
for the study of cross-linked collagen strucHackman, R. H. 1984. Cuticle biochemistry. In J.
tures. Trans. N.Y. Acad. Sci. Series II, 34:694Bereiter-Hahn, A. G. Matoltsy, and K. S. Rich708.
ards (eds.), Biology of the integument. 1. Invertebrates,
Liem, K. F. 1981. Larvae of air-breathing fishes as
pp. 583—610. Springer-Verlag, New York.
countercurrent flow devices in hypoxic environHadley, N. F. 1980. Surface waxes and integumenments. Science 211:1177-1179.
tary permeability. Amer. Sci. 68:546-553.
Lillywhite, H. B. 1971. Thermal modulation of cutaHadley, N. F. 1981. Cuticular lipids of terrestrial
neous mucus discharge as a determinant of evapplants and arthropods: A comparison of their
orative water loss in the frog, Rana catesbeiana.
structure, composition, and waterproofing funcZ. Vergl. Physiol. 73:84-104.
tion. Biol. Rev. 56:23-47.
Lillywhite, H. B. 1975. Physiological correlates of
Hadley, N. F. 1984. Cuticle: Ecological significance.
basking in amphibians. Comp. Biochem. Physiol.
In J. Bereiter-Hahn, A. G. Matoltsy, and K. S.
52A:323-330.
Richards (eds.), Biology of the integument. 1. Invertebrates, pp. 685—693. Springer-Verlag, New York. Lillywhite, H. B. and P. F. A. Maderson. 1982. Skin
structure and permeability. In C. Gans and F. H.
Hama, K. 1969. The fine structure of some blood
Pough (eds.), Biology of the Reptilia, Vol. 12 Physvessels of the earthworm, Eisenia foetida. J. Bioiology C, Physiological ecology, pp. 397-442. Acaphys. Biochem. Cytol. 7:717-724.
demic Press, New York.
Hauser, H., A. Darke, and M. C. Phillips. 1976. Ion
MacDougall.J. D. B. and M. McCabe. 1967. Diffubinding to phospholipids: Interaction of calcium
sion coefficient of oxygen through tissues. Nature
with phosphatidylserine. Eur. J. Biochem. 62:335215:1173-1174.
344.
Machin, J. 1964a. The evaporation of water from
Hauser, H. and M. C. Phillips. 1979. Interactions of
Helix aspersa. I. The nature of the evaporating
the polar groups of phospholipid bilayer memsurface. J. Exp. Biol. 41:759-769.
branes. Prog. Surf. Membr. Sci. 13:297-413.
Hevesy, G., E. Hofer, and A. Krogh. 1935. The per- Machin, J. 19646. The evaporation of water from
Helix aspersa. II. Measurement of air flow and the
meability of the skin of frogs to water as deter-
962
H . B. LlLLYWHITE AND P. F. A. MADERSON
diffusion of water vapor. J. Exp. Biol. 41:771781.
Machin,J. 1977. Role of integument in molluscs. In
B. L. Gupta, R. B. Moreton, J. J. Oschman, and
B. J. Wall (eds.), Transport of ions and water in
animals, pp. 735-762. Academic Press, New York.
Maderson, P. F. A. 1972. When? Why? and How?:
Some speculations on the evolution of the vertebrate integument. Amer. Zool. 12:159-171.
Mahany, T. M. and R. H. Parsons. 1978. Circulatory
effects on osmotic water exchange in Rana pipiens. Amer.J. Physiol. 234:R172-R177.
Malvin, G. M. 1988. Microvascular regulation of
cutaneous gas exchange in amphibians. Amer.
Zool. 28:999-1007.
Mangum, C. P., B. R. McMahon, P. L. deFur, and M.
G. Wheatly. 1985. Gas exchange, acid-base balance, and the oxygen supply to the tissues during
a molt of the blue crab Callinectes sapidus. J. Crustacean Biol. 5:188-206.
Marshall, W. S. 1985. Paracellular ion transport in
trout opercular epithelium models osmoregulatory effects of acid precipitation. Can.J. Zool. 63:
1816-1822.
Matoltsy, A. G. 1984. Structure and function of the
mammalian epidermis. In J. Bereiter-Hahn, A.
G. Matoltsy, and K. S. Richards (eds.), Biology
of the Integument. 2. Vertebrates, pp. 255—271.
Springer-Verlag, New York.
Mauro, A. 1957. Nature of solvent transfer in osmosis. Science 126:252-253.
McClanahan, L. L., J. N. Stinner, and V. H. Shoemaker. 1978. Skin lipids, water loss, and energy
metabolism in a South American tree frog (Phyllomedusa sauvagei). Physiol. Zool. 51:179-187.
Mittal, A. K. and J. S. Datta Munshi. 1971. A comparative study of the structure of the skin of certain air-breathing fresh-water teleosts. J. Zool.
163:515-532.
Neumcke, B. 1971. Diffusion polarization at lipid
bilayer membranes in the presence of a homogeneous chemical reaction in the solutions. T.I.T.
J. Life Sci. 1:85-90.
Paganelli, C. V., R. A. Ackerman, and H. Rahn. 1978.
The avian egg: In-vivo conductances to oxygen,
carbon dioxide, and water vapor in late development. In]. Piiper (ed.), Respiratory function in
birds, adult and embryonic, pp. 212-218. SpringerVerlag, Berlin.
Piiper, J. 1988. Models for cutaneous gas exchange
and transport. Amer. Zool. 28:963-972.
Quinlan.M.C. andN. F. Hadley. 1982 A new system
for concurrent measurement of respiration and
water loss in arthropods. J. Exp. Zool. 222:255263.
Rabl, H. 1931. Integument der Anaminier. In L.
Bolk, E. Gdppert, E. Kallius, and W. Lubosch
(eds.), Handbuch der vergleichenden Anatomic der
Wirbelthiere, Vol. 1, pp. 271-374. Urban and
Schwarzenberg, Berlin.
Randall, D. J. 1970. The circulatory system. In W.
S. Hoar and D. J. Randall (eds.), Fish physiology,
Vol. 4, pp. 133-172. Academic Press, New York.
Rieger, R. M. 1984. Evolution of the cuticle in the
lower Eumetazoa. In J. Bereiter-Hahn, A. G.
Matoltsy, and K. S. Richards (eds.), Biology of the
integument. 1. Invertebrates, pp. 389—399. SpringerVerlag, New York.
Schafer.J. A.andT. E. Andreoli. 1972. Water transport in biological and artificial membranes. Arch.
Intern. Med. 129:279-292.
Shick, J. M., W. I. Brown, E. G. Dolliver, and S. R.
Kayar. 1979. Oxygen uptake in sea anemones:
Effects of expansion, contraction, and exposure
to air and the limitations of diffusion. Physiol.
Zool. 52:50-62.
Simkiss, K. and K. M. Wilbur. 1977. The molluscan
epidermis and its secretions. Symp. Zool. Soc.
London 39:35-76.
Stokes, G. D. and W. A. Dunson. 1982. Permeability
and channel structure of reptilian skin. Amer.J.
Physiol. 242:F681-F689.
Ultsch.G. R.andG. Gros. 1979. Mucus as a diffusion
barrier to oxygen: A possible role in oxygen
uptake at low pH in carp (Cyprinus carpio) gills.
Comp. Biochem. Physiol. 62A:685-689.
Vincent,). F. V. andj. E. Hillerton. 1979. The tanning of insect cuticle—a critical review and a
revised mechanism. J. Insect Physiol. 25:653-658.
Vogel, W. O. P. 1985. Systemic vascular anastomoses, primary and secondary vessels in fish, and
the phylogeny of lymphatics. In K. Johansen and
W. W. Burggren (eds.), Cardiovascular shunts,
Alfred Benzon Symposium 21, pp. 143-151.
Munksgaard, Copenhagen.
Wertz, P. W. 1986. Lipids of keratinizing tissues. In
J. Bereiter-Hahn, A. G. Matoltsy, and K. S. Richards (eds.), Biology of the integument. 2. Vertebrates,
pp. 815-823. Springer-Verlag, New York.
Whitear, M. 1977. A functional comparison between
the epidermis of fish and of amphibians. Symp.
Zool. Soc. London 39:291-313.
Wilson, R. A. 1968. An investigation into the mucus
produced by Lvnnaea truncatula, the snail host of
Fasciola hepatica. Comp. Biochem. Physiol. 24:
629-633.
Wright, E. M., A. P. Smulders, and J. M. Tormey.
1972. The role of the lateral intercellular spaces
and solute polarization effects in the passive flow
of water across the rabbit gallbladder. J. Membrane Biol. 7:198-219.