AMER. ZOOL., 28:761-773 (1988)
Endocrine Aspects of Homeostasis1
JANE C. KALTENBACH
Department of Biological Sciences, Mount Holyoke College,
South Hadley, Massachusetts 01075
SYNOPSIS. Recent findings in endocrine research have greatly increased our understanding of the relationship between hormones and homeostasis. The present paper reviews
selected major advances in such areas as neuropeptides, peptide biosynthesis in endocrine
and neuronal cells, peptide receptors and intracellular pathways in target cells, "new"
peptide hormones, and evolutionary considerations of peptide hormones. Further understanding of hormone interactions and of relationships between nervous, endocrine, and
immune systems has added to the growing complexity of the mechanisms of fine tuning
and regulating our internal environment. Moreover, discovery of the same or similar
peptides throughout the course of evolution, i.e., from unicellular organisms through
vertebrates, has led to a new unifying theory of intercellular communication. Endocrine
aspects of homeostasis is an expanding and exciting field of biology.
INTRODUCTION
"Endocrine aspects of homeostasis" is a
broad title and one which is becoming even
broader, as strict, traditional definitions of
these terms are modified or replaced by
more comprehensive ones, and as the complexities of hormonal interrelationships are
better understood.
Maintenance of a relatively stable internal environment within the body enables
cells to function under optimal conditions,
thereby enabling organisms to be independent of the sometimes widely fluctuating
factors outside of the body. The concept
of a constant internal environment, first
formulated by the French physiologist
Claude Bernard in the late 1800s, was later
further developed and given the name
homeostasis by Walter Cannon. The somewhat newer term homeokinesis reflects the
dynamic nature of the process as well as its
applications to all levels of life from molecules to organisms to communities. Whatever its name, the concept ranks among
the most important in biology.
The endocrine system plays a major role
in regulating the stability of the cellular
environment. Can you think of any homeostatic mechanism that is not controlled at
least in part by endocrine glands and their
hormones? Probably not. Maintenance of
temperature, ions, water, etc. are all held
within optimal ranges by hormone actions
or interactions. Historically, a hormone has
been defined as a glandular secretion which
is transported in the blood and evokes
responses in specific target organs. Should
the term now be extended to include other
newly discovered substances which in one
way or another do not quite fit this definition (e.g., neuropeptides, paracrines,
growth factors)? One may even ask if there
is any tissue in the body which does not
secrete at least one hormone and, on the
other hand, which does not also function
as a target organ.
Recent advances in endocrine research
have greatly enhanced our understanding
of the classical relationship between the
endocrine system and homeostasis (Fig. 1).
The present paper focuses on selected
advances in peptide research which contribute new perspectives to our understanding of the control and fine-tuning of
hormone concentrations in the blood, and
hence the homeostatic mechanisms which
they in turn regulate.
NEUROPEPTIDES
Neurohormone is an appropriate term
for peptides which are synthesized in neurons (rather than in glandular cells), are
carried in the circulation, and evoke
responses in target organs (such as the kidney, uterus, and even endocrine glands).
1
From the Symposium on Science as a Way of KnowThe first peptides isolated from the brain
ing—Form and Function presented at the Annual
Meeting of the American Society of Zoologists, 27- were the two mammalian neurohypophy30 December 1987, at New Orleans, Louisiana.
seal hormones, vasopressin (or antidiuretic
761
762
JANE C. KALTENBACH
TRH (thyrotropin-releasing hormone),
LHRH (luteinizing hormone-releasing
hormone), and somatostatin (growth hormone-inhibiting hormone). The prize was
Feedback
Hormones
also shared with Rosalyn Yalow for development of the RIA (radioimmunoassay), a
technique which for the first time made
possible the measurement of minute quanTarget Organs
tities of hormones in blood and tissue
Homeost at i c extracts. Since then, CRH (corticotropinMechani sms
releasing hormone) and GHRH (growth
hormone-releasing hormone) have also
\
Internal Environment
been synthesized (Table 1). Such regulatory peptide hormones are released from
FIG. 1. Endocrine control of homeostasis.
hypothalmic neurons into the hypophyseal
portal system, the vascular link to the antehormone, ADH) and oxytocin. These are rior pituitary; there they affect the release
synthesized in nerve cell bodies in specific of pituitary hormones into the circulation,
regions of the hypothalamus, released from and directly or indirectly regulate homeoaxon endings into capillaries in the poste- static mechanisms related to metabolism,
rior pituitary (neurohypophysis), and hence reproduction, growth, stress, etc. Such facto distant targets, for regulation of blood tors as negative feedback loops, the biopressure, water balance, uterine contrac- logical clock which governs endocrine
tion, etc. Characterization and synthesis of cycles, and the episodic or pulsatile nature
these neurohormones was a feat for which of neurohormone and hormone release add
Vincent duVigneaud was awarded a Nobel to the complexities of the neuroendocrine
Prize in 1958. This was an important land- system.
mark in the now burgeoning field of neuReleasing and inhibiting hormones have
roendocrinology.
been demonstrated by immunohistochemA few years later a highly competitive ical techniques not only within specific
race {e.g., review: Crapo, 1985), for iden- locations in the hypothalamus, but also in
tification and synthesis of hypothalmic hor- neurons throughout the central nervous
mones which regulate the anterior pitu- system (CNS). Such a wide distribution sugitary resulted in another Nobel Prize gests that hypothalamic peptides function
(1977), this time for Andrew Schally and not only as neurohormones but also as neuRoger Guillemin; they independently rotransmitters or neuromodulators. Single
characterized and synthesized the peptides peptides with dual roles!
Endocri ne Glands
TABLE 1.
Peptide hormones of the mammalian hypothalamus.
Hormone
Neurohypophyseal hormones
Oxytocin
Vasopressin or antidiuretic hormone, ADH
Releasing and inhibiting hormones
Thyrotropin-releasing hormone, TRH
Luteinizing hormone-releasing hormone, LHRH
Growth hormone-inhibiting hormone, somatostatin
Corticotropin-releasing hormone, CRH
Growth hormone-releasing hormone, GHRH
Prolactin-releasing hormone, PRH
Prolactin-inhibiting hormone, PIH
No. amino acids
9aa
9aa
3 aa
10 aa
14 aa
41 aa
44 aa
Synthesis
1953 du Vigneaud
1954 du Vigneaud
1969
1971
1973
1981
1982
Schally and Guilleman
Schally and Guilleman
Schally and Guilleman
Vale
Vale and Guilleman
763
ENDOCRINE ASPECTS OF HOMEOSTASIS
Within the last decade classical anterior
pituitary hormones, gastrointestinal hormones, and others such as calcitonin, and
angiotensin II have been detected by means
of different approaches (including molecular probes) in specific regions of the brain
(reviews: Krieger and Liotta, 1979; Snyder, 1980; Krieger, 1983). Evidence now
indicates that some are actually synthesized
within the CNS, although it has not been
established whether or not one neuron can
produce and secrete more than one peptide. Possible roles include neurotransmission or neuromodulation, coordination of
responses to stimuli, and regulation of a
variety of homeostatic and behavioral systems related to feeding, temperature, blood
pressure, learning and memory, pain, and
stress (Krieger, 1983). In fact such systems
are undoubtedly regulated by multiple
neuropeptides.
Further complicating the picture are
observations of colocalization of neuropeptides and classical neurotransmitters
(e.g., acetylcholine) within the same neurons (review: Hokfelt et al., 1987). Still to
be answered are such questions as how
common this situation is in the brain of
mammals and other vertebrates, and
whether or not it is of physiological significance. Informational traffic in synapses is
certainly more complex than previously
believed. The discoveries of ever increasing numbers of peptides in the brain have
revolutionized our ideas of neuronal messengers and have placed neuropeptides in
a central position in the direct or indirect
regulation of homeostasis.
PEPTIDE BIOSYNTHESIS
The mechanisms by which regulatory
peptides are synthesized are similar in both
endocrine and neuronal cells. Biosynthesis
of peptide and neuropeptide hormones
involves transcription and translation of
large, inactive precursor molecules, followed by posttranslational processing and
release of biologically active peptide hormones. The morphological pathway from
the site of precursor synthesis on ribosomes to hormone exocytosis at the cell
membrane is also similar in all peptidesecreting cells (Fig. 2). Alterations in any
pREfRDHORfVONE
EPRl
HORMONE
FIG. 2. Peptide biosynthesis: Posttranslational processing and its intracellular pathway.
of the biochemical steps along this intracellular pathway could affect concentrations of peptide hormones in the circulation and thus regulation of the constancy
of the internal environment.
Synthesis of the precursor molecules may
be influenced by rates of transcription and
translation and mechanisms controlling
gene expression. Although one gene may
code for one precursor, in some cases,
alternative processing of the same mRNA
results in two precursors and therefore different active peptides. In addition, enzymatic cleavage of a precursor molecule at
different sites will lead to release of different peptide hormones (review: Solcia et al.,
1987).
After translation on ribosomes, precursors are directed across the endoplasmic
reticulum (ER) membrane and into the ER
cisterna, according to the signal hypothesis
(Blobel and Dobberstein, 1975). Many
details of this translocation process have
been worked out within the last ten years.
A large precursor, called a preprohormone, has at its amino terminal a signal
sequence of from 16 to 30 amino acid residues, the core of which is hydrophobic.
As the preprohormone is being formed on
free ribosomes, a signal recognition particle, SRP, (which recognizes the signal
sequence) binds to the ribosome and then
interacts with an SRP receptor (docking
protein) in the ER membrane. Latest indi-
764
JANE C. KALTENBACH
FIG. 3. Signal hypothesis: Translocation of a peptide
to and across an ER membrane. R, ribosome; S, signal;
SRP, signal recognition particle; SRP REC, signal recognition particle receptor; SSR, signal sequence
receptor.
cations are that after release from the SRP
complex, the signal interacts with a newly
discovered second membrane receptor, a
signal sequence receptor (SSR), believed to
be associated with a translocation pore
(Wiedmann et al, 1987). By such mechanisms the signal is responsible for directing
the nascent peptide from a ribosome to and
across the ER membrane, (Fig. 3). The
intricacies of this process may enhance targeting specificity and/or prevent folding
of the peptide prior to translocation.
After entering the ER cisterna, the signal is rapidly removed by a peptidase. The
resultant prohormone is transferred to the
Golgi for possible modification (sulfation,
phosphorylation, glycosylation, etc.), as well
as sorting and packaging into regulated
secretory vacuoles. Sorting of various proteins into different vacuoles may be directed
by sorting domains on the peptide (Moore,
1987). It is within these dynamic organelles
(no longer considered to be mere storage
sites) that the prohormone is cleaved by
endopeptidases at specific sites, i.e., at one
or two basic amino acid residues (arginine
and/or lysine); the latter are then removed
by carboxy or amino-peptidases (reviews:
Andrews^ al, 1987; Marx, 1987a).
Such posttranslational cleavage by prohormone cleaving enzymes (currently being
identified) may yield various types of peptides (Fig. 4) including, e.g., a single linear chain, parathyroid hormone (PTH)
(review: Potts et al, 1980), two S-linked
chains, insulin (review: Tager et al, 1980),
more than one active peptide adrenocorticotropic hormone (ACTH), melanophore-stimulating hormone (MSH) (reviews: Andrews et al, 1987; Costa et al,
1987), and multiple copies of the same peptide, TRH, (Lechan et al, 1986). (Further
processing may occur outside of the cell,
e.g., cleavage of PTH [84 amino acids] to
a more active 34 amino acid chain, deiodination of T 4 to the more active T3.)
Upon receipt of a proper stimulus, peptide hormones are released from cells by
exocytosis. Secretory vacuoles move and
fuse to the cell membrane; a fusion pore
(the size of a single gap junction channel)
forms and then dilates, thereby releasing
the hormone to the outside of the cell
(Breckenridge and Aimers, 1987). On the
other hand, if the secretory vacuoles should
be in excess (e.g., in pituitary mammotrophs at the end of nursing), their hormone may be enzymatically degraded upon
fusion of the vacuoles to lysosomes.
The interactions involved in targeting
and translocation of hormone precursors
across ER membranes, the nature and
reactions of prohormone cleaving enzymes,
and the mechanisms of exocytosis are currently under intense investigation. Integration of future results into our present
hypotheses will surely enhance our understanding of hormone regulated homeostasis.
PEPTIDE RECEPTORS AND
INTRACELLULAR MESSENGERS
Mechanisms by which peptide hormones
bring about physiological responses in target cells and thereby influence the surrounding extracellular environment is a
topic of enormous interest to present day
endocrinologists. Peptide hormones rather
than entering their target cells bind with
765
ENDOCRINE ASPECTS OF HOMEOSTASIS
Si
PTH
ACTH
Si
Si
1TRH1
1
MSH
MSH
MSHENK
FIG. 4. Peptides resulting from posttranslational cleavage, (a) one linear peptide chain, PTH; (b) two S-linked
peptide chains, INS; (c) multiple peptides, ACTH, LPT, END, MSH, ENK; (d) multiple copies of one peptide,
TRH. Si, signal; INS, insulin; LTP, lipotropin; END, endophin; ENK, enkephalin.
high affinity to specific protein receptors
in the outer cell membrane. A chain of
enzymatic reactions triggered by this event
is responsible for the appropriate physiological response.
Results of cloning and sequence analysis
of genes for several receptors have indicated that despite differences among peptide hormones, receptors are similar in
structure and size, as well as in number and
sequence of amino acid residues, especially
in the hydrophobic, helical segments
located within the cell membrane (Fig. 5).
Moreover, the genes themselves have no
introns, and are probably members of the
same family and descendents of a common
ancestor gene (review: Marx, 1987c).
Regulation of receptor numbers is of
critical importance and depends upon
receptor synthesis and degradation, so
called up and down regulation (review:
Kahn, 1976). As concentrations of most
hormones increase, there is a loss of their
own receptors, but in other cases (e.g., prolactin) the opposite is true and the number
of receptors increase. Moreover, there are
a number of examples of heterologous regulation of receptors, e.g., with FSH stimulation, the number of LH receptors in the
ovary is up regulated. As new information
accumulates, the complexities and impor-
tance of factors which control receptors
are becoming more evident.
A related and expanding field of research
is centered upon G proteins (guanosine triphosphate or GTP binding proteins) which
act as intermediaries, coupling receptor
molecules to enzymatic pathways within the
cell (review: Vaughan, 1987). In fact, this
protein is probably involved in most receptor-linked cellular responses. Binding of a
hormone, a first messenger, to its receptor
activates a G protein which in turn functions as an on-off switch for enzymes such
as adenylate cyclase responsible for formation of cAMP (Fig. 6). This is the now
classical second messenger within the cell
which earned a Nobel Prize for Earl Sutherland in 1971. G proteins have been structurally analysed (three subunits), and some
have been isolated. In addition to Gs and
Gi which activate and inhibit adenylate
cyclase, new types include Gt (visual transduction), Go (function not established), Ge
(control of exocytosis), Gp (activation of
phospholipase C) Gk (activation of potassium channels). There are undoubtably
many more. Moreover, dual pathways may
be used by Gs to regulate Ca + + channels,
i.e., indirectly by means of cAMP and kinases and directly without cytoplasmic
mediators (Yatani et al, 1987). Possible
766
JANE C. KALTENBACH
FIG. 5. Receptor structure: /3s-adrenergic receptor,
a protein with seven helical segments of hydrophobic
amino acids in the cell membrane and three loops of
hydrophilic amino acids on each side of the membrane.
interactions of G proteins with each other
and with various second messenger systems
provide exciting new avenues to explore.
Many peptide hormones evoke responses
in target tissue through intracellular pathways involving ATP and the formation of
cAMP (or cGMP) (review: Sutherland,
1972). This second messenger activates
cAMP (or cGMP) dependent protein kinase
needed to start phosphorylation of other
proteins (Fig. 6). A series of such reactions
forms a cascade of amplification of the
original hormone signal.
In the last few years interest has focused
on what is proving to be another major
pathway, the polyphosphoinositide (PI)
system (review: Joseph, 1985). Hormone
binding to a G linked receptor activates
phospholipase C which splits a membrane
lipid (PIP2 or polyphosphatidyl-4,5biphosphate) thereby releasing two second
messengers, inositol triphosphate (IP3) and
1,2 diacylglycerol (DAG) (Fig. 7). The Ca+ +
released by IP3 from intracellular storage
sites plays a direct role in many cellular
responses and together with DAG stimulates protein kinase C, another important
regulator of cellular enzymatic activities
(reviews: Berridge and Irvine, 1984;
Majerus etal, 1985; Berridge, 1987; Marx,
19876). Current research is showing the
importance of the PI system, including its
functions within the brain related to regulation of ion channels, neurotransmission, and memory and learning.
The number of identified protein kinases, i.e., enzymes with ability to phosphorylate other proteins, has increased
from a few in 1980 to almost a hundred
today; catalytic domains have been found
to be similar in all of them (review: Hunter,
1987). Why so many kinases? Why so many
in the brain? What types of metabolic pathways do they regulate? How many substrates are there in a cell? These are just a
few of the many questions now being investigated.
The complexity and interactions of second messenger systems within many target
cells are illustrated by arachidonic acid
metabolites or eiocosanoids (a Nobel Prize,
this time for Bergstrom, Samuelsson, and
PEPTIDE HORMONE
1ST MESSENGER
CELL
MEMBRANE
I Gs ProteTTn
adeny]_ate cyclase
ATP'
CYTOPLASM
OF
TARGET CELL
cAMP
2nd MESSENGER
5' AMP
INACTIVE
cAMP DEPENDENT
PROTEIN KINASE
PROTEIN
PHOSPHORYLATIONS
RESPONSE
FIG. 6. cAMP receptor system.
767
ENDOCRINE ASPECTS OF HOMEOSTASIS
PEPTIDE HORMONE
1 S T MESSENGER
PROTEINICELL
\
MEMBRANE 1
PHOSP
OSPHOL IPASEC
HEJ
PIP2-
CYTOPLASM
OF
TARGET CELL
ITP
2nd messenger
DAG
2nd messenger
Ca+ +
(From Storage s i t e s ) .
PROTEIN KINASE C
PROTEIN
PHOSPHORYLATIONS
GENE
ACTIVATION
ION CHANNEL
REGULATION
lESPONSES*^
FIG. 7. Polyphosphoinositide (PI) receptor system.
Vane, 1982). Within the cell membrane
phospholipases release arachidonic acid
from phospholipids (reviews: Nelson et al.,
1982; Bevan and Wood, 1987; Samuelsson
et al., 1987). Depending upon the tissue
and the hormone message, arachidonic acid
may be processed by either the cyclooxygenase or lipoxygenase pathways; the first
yields prostaglandins, thromboxanes, and
prostacyclins and the second leukotrienes
and lipoxins. Prostaglandins and thromboxanes may be considered as second messengers, intermediaries in the activation of
adenylate or guanylate cyclases (which then
may be considered as third messengers)
(Fig. 8). On the other hand, prostacyclins,
and leukotrienes, and probably lipoxins,
are released from the cell and as good hormones, travel in the blood to target cells,
where they induce responses through the
nucleotide cyclases. The diversity of physiological functions in which such compounds are involved is huge and will remain
a topic for another discussion.
A more speculative intracellular pathway involves dissociation of G proteins into
leukotrienesl | pr ost acy cl i nsl
(hormones)
\
^
(hormones)
Hormone
(1 st messenger^
PHOSPHO
PHOSPHOLIPIDS
CYTOPLASM
OF
TARGET CELL
prostagl andl ns &
t hromboxanes
cAMPI
3rd messenger
4P r o t e l n Kl nase
Protein
I
Phosphor ylat I ons 4.
Response
FIG. 8. Arachidonic acid metabolites as hormones and as second messengers.
768
JANE C. KALTENBACH
programmable messengers. According to
this idea hormone-binding to receptors
causes G proteins to release their alpha
subunits (primary messengers) into the
cytoplasm where they may be modified or
programmed in different ways, thus making possible multiple cellular responses
(Rodbell, 1985).
The finding of LDL (low density lipoprotein) as well as peptide hormones and
receptors within the cytoplasm of target
cells led to the theory of receptor-mediated
endocytosis (reviews: Goldstein etal., 1979,
1985; Dautry-Varsat and Lodish, 1984;
Carpentier^a/., 1986). For example, insulin-receptor complexes in the cell membrane cluster in pits coated with clathrin,
and then are internalized by invagination
and formation of coated vesicles. Upon loss
of the clathrin coat, several vesicles fuse
and form an endosome, in which the hormone-receptor bonds are broken. This
permits the receptors to be recycled and
the hormone to be degraded by lysosomal
enzymes. Whether the insulin ever plays a
functional role within the cytoplasm or
whether the main purpose of this system is
to control numbers of receptors or to stop
hormone-triggered reactions has not been
established. Such possibilities are of importance with respect to glucose homeostasis.
Clarification of biochemical aspects of
intracellular systems within target cells
should yield exciting new insights into the
mechanisms by which peptides regulate
homeostatic mechanisms.
NEW PEPTIDE HORMONES
Atrial natriuretic factor (ANF)
Certain peptides that differ in one way
or another from the classical definition of
a hormone have now been added to the
repertoire of hormones that regulate
homeostatic mechanisms. ANF is one such
peptide; it is synthesized, not by glandular
tissue, but rather by cardiac muscle cells,
primarily within the atria. As its name
implies, it promotes excretion of sodium
and water from the body, and by interacting with other hormone systems plays a
role in fine-tuning blood pressure and volume homeostasis. Since 1980 a tremendous amount of information has been
reported about its regulation, biosynthesis,
biochemical identity, function, and even
mechanisms of action within target cells
(reviews: Anderson and Bloom, 1986; Cantin and Genest, 1986). As is true for other
peptides, it is synthesized as a large preprohormone (in the rat, 152 amino acids)
which includes at the N terminal a 24 amino acid signal and at the C terminal a 28
amino acid sequence which becomes the
active circulating hormone. ANF tends to
decrease blood pressure in several different ways, e.g., relaxing arterial smooth
muscle, inhibiting secretion of aldosterone
and renin, stimulating glomerular permeability and decreasing reabsorption of
sodium from kidney tubules, as well as acting on a number of sites in the brain that
are associated with blood pressure. Moreover, the recent finding of ANF in the choroid plexus epithelium of the brain suggests that this peptide influences the
secretion of cerebral spinal fluid and hence
is an important factor in water regulation
within the central nervous system (Steardo
and Nathanson, 1987). Receptors in the
choroid are associated with guanylate
cyclase and GMP production as are ANF
receptors in the periphery of the body.
Certainly, control of homeostatic mechanisms regulating blood volume and blood
pressure are complex!
Hematologic growth factors
These glycoprotein hormones are so
named because of their ability to stimulate
blood cell production in the bone marrow.
Erythropoietin (Ep) stimulates production
of erythrocytes by causing differentiation
and hemoglobin production in committed
erythroid cells. Four colony-stimulating
factors (CSFs) increase numbers of leukocytes by stimulating proliferation or cloning of committed progenitor cells (they
form colonies in semisolid agar, hence the
name). None of them are synthesized in
the usual endocrine glands, i.e., Ep in the
kidney and CSFs in multiple cell types in
all tissues (e.g., endothelial cells, fibroblasts, macrophages, lymphocytes). Ep has
been known for some years, and its regulation and function are quite well known.
The general functions of the various types
ENDOCRINE ASPECTS OF HOMEOSTASIS
769
of CSFs are indicated by their names: GM- the immune system. Lymphokines act on
CSF, granulocyte macrophage-colony- the CNS, where, for example, they activate
stimulating factor, which has been well opioid receptors and stimulate glial cells
characterized; G-CSF, granulocyte-colony- (review: Morely et al., 1987). Interleukin-1
stimulating factor; M-CSF, macrophage- (IL-1) from monocytes stimulates ACTH
colony-stimulating factor; Multi-CSF (or secretion either directly by action on pituinterleukin-3), which acts on stem cells very itary cells or indirectly by stimulation of
early in their differentiation and therefore CRH (Lumpkin, 1987). Moreover, pepis able to stimulate granulocytes and mac- tides from the thymus mediate response in
rophages as well as T cells (review: Kolata, the endocrine and neural systems. For
1987). Hematologic growth factors are now example, thymosin |84, by acting on the
subject to intense research efforts. Recently hypothalamus, causes the pituitary to
genes for human Ep and the four CSFs release LH and so stimulates other endohave been cloned, and receptor numbers/ crine glands; thymosin 5 stimulates release
cell and molecular weights of CSFs as well of ACTH and endorphins from cultured
as their responsiveness to environmental pituitary cells, and thymosins and lymfactors (e.g., antigens) have been deter- phokines directly influence release of the
mined (reviews: Metcalf, 1985; Clark and neurotransmitter noradrenaline in the
Kamen, 1987; Kolata, 1987). The impor- brain (reviews: Hall and Goldstein, 1986;
tance of Ep and CSFs is assured with respect Morely et al., 1987). Neuro-endocrine—
to regulation of blood cell homeostasis and immune interactions are just beginning to
to their potential in clinical situations, unfold as are the ways in which they proespecially those involving infection and tect the body and maintain the homeostatic
immune reactions.
mechanisms of the defense system. Details
of these interactions and of intracellular
pathways in the target cells remain to be
Immunotransmitters
Peptides which are secreted by cells worked out in the future, along with charwithin the immune system and act upon acterization of the genetic determinants of
targets within the same system have been the immunotransmitters and the cell
called immunotransmitters. They are also receptors. A new integrative science is on
considered as hormones secreted, not by the way!
endocrine glands, but by circulating
EVOLUTION OF PEPTIDE HORMONES
immune cells, which are important in conUnifying
theory of intercellular communication
trolling the homeostatic mechanisms of the
body's defense system. Until recently,
Hormonal and neuronal peptides idenimmunotransmitters were thought to con- tical, or very similar chemically, to those
sist of the many lymphokines and cytokines in vertebrates have been found in inverwhich influence the body's immune tebrates, plants, and unicellular organisms
responses. However, new reports indicate including protozoans, yeast, and bacteria.
that lymphocytes and macrophages also Such findings plus the presence of approrelease peptides once thought to be priate receptors is an indication that pepsecreted only by the pituitary and the brain, tides act as chemical messengers even in
e.g., ACTH, endorphins, and enkephalins, primitive organisms. Many of these horand possibly even CRH, TSH, growth hor- mone-like messengers in unicellular ormone, chorionic gonadotrophin, FSH and ganisms are involved with feeding or reLH; in turn many neuropeptides bind to productive activities (review: Roth and
lymphocytes and monocytes as targets LeRoith, 1987). Examples include ACTH
(reviews: Hall and Goldstein, 1986; Payan and endorphins in protozoans and an
et al., 1986; Morely et al., 1987). Not only LHRH-like peptide, which is involved in
have the known numbers of peptides mating of yeast cells. Relevant receptors
secreted by immune cells increased in the are also similar to those in vertebrates
last few years, but so have the numbers of (LeRoith et al., 1986). Primitive chemical
target cells both outside as well as inside communication, like that in vertebrate sys-
770
JANE C. KALTENBACH
terns, comprises secretory cells, chemical
substances (hormones) carried in the
medium (extracellular fluids), and responding cells (targets). However, in unicellular organisms the secretory cells and
the target cells are in separate organisms
(similar to pheromones in multicellular
organisms). Despite evolving specialization
and complexity of secretory and target cells,
theory now has it that during the long
course of evolution the chemical structures
of the messengers have changed very little,
i.e., they have been highly conserved; this
is the new "unifying theory of intercellular
communication" (reviews: Kolata, 1982;
Roth etal, 1982; Roth and LeRoith, 1987).
Moreover, it is now believed that both
endocrine and nervous systems are descended independently from a common
ancestral system (review: Roth and
LeRoith, 1987). Thus many peptides probably present in ancestral organisms act both
as hormones and neurotransmitters in vertebrates. The overlap between chemical
messengers in vertebrate endocrine and
exocrine glands may be similarly explained.
A unitary origin of these two types of glands
may account for the presence of hormonal
(or neurohormonal) peptides, such as prolactin, gastrin, somatostatin, TRH, in various exocrine secretions (review: Roth et
al., 1982). We still do not know the functions of insulin in spinach and of TRH in
alfalfa or for that matter in glands of
amphibian skin. This presents exciting
areas for investigation. How early in evolutionary history were such compounds
involved in homeostatic regulation?
FIG. 9. Rana clamitans tadpole showing local resorption of the dorsal tail fin in the region of a thyroxine
implant, indicating direct hormonal action upon
peripheral tissues. Unpublished photomicrograph of
Jane C. Kaltenbach.
promoted by direct action of thyroxine upon
peripheral target tissues. Since metamorphic changes were accelerated locally in
tissues adjacent to thyroxine implants we
concluded that, indeed, such was the case
(Fig. 9). In addition, local metamorphic
events induced by certain thyroxine analogue implants indicated that specific analogues act directly upon peripheral tissues;
chemical changes in the analogues, if any,
would have occurred locally within the target tissues (review: Kaltenbach, 1968). To
determine if other hormones interact with
thyroxine, hormone implants and cultures
of tail-tips in hormone solutions were used;
results indicated that hydrocortisone, cortisone, and deoxycorticosterone enhance
the metamorphosing activity of thyroxine
and do so by shortening its latent period
(Kaltenbach, 1958, 1985).
Moreover, serum thyroxine concentrations have been measured by RIA. Hormone levels peak during metamorphic climax, but thyroxine levels were below the
limits of the assay in early-stage tadpoles
(Mondou and Kaltenbach, 1979). However, indirect immunofluorescent staining
Amphibian metamorphosis
of blood smears from such tadpoles indiWith such thoughts in mind, I should cated the presence of some thyroxine, not
like to conclude this discussion by men- only in the serum, but also in the red blood
tioning my own research on endocrine cells (Piotrowski and Kaltenbach, 1985).
aspects of amphibian metamorphosis. By a Fluorescence indicative of thyroxine was
slight stretch of the imagination metamor- especially intense in the nuclei of red blood
phosis may be considered under the term cells of climax tadpoles and frogs (Stieff
homeostasis. After all if the internal envi- and Kaltenbach, 1986); in addition, recepronment of the developing tadpole were tors for thyroid hormone have been found
not carefully regulated, would a healthy in amphibian erythrocyte nuclei (Galton,
1984, 1985; Moriya et al, 1984; Galton
frog result?
and
St. Germain, 1985). Yet the function
Metamorphic events involving both form
of
thyroxine
in such target cells is still open
and function are dependent upon thyroid
hormone. We first asked if such events were to question.
ENDOCRINE ASPECTS OF HOMEOSTASIS
FIG. 10. Frozen sections of Rana pipiens frog skin;
fixed in acrolein and stained for TRH with the silverenhanced immunogold method. Unpublished photomicrographs of Beek Yoke Chin, Mount Holyoke
College, (a) gold particles (black), indicative of TRH,
are visible in a serous (granular) gland; (b) control
section with non-immune serum as replacement for
the primary antiserum. Gold particles are not detectable in the gland.
Currently we have turned our attention
to a possible relationship between TRH,
the pituitary-thyroid axis, and metamorphosis. TRH injections, under a wide range
of conditions, had no effect on tadpole
growth or metamorphosis, suggesting
that TRH does not stimulate release of
either prolactin (a tadpole growth hormone) or thyroid hormone (Kaltenbach and
Mimnagh, 1986), despite such effects in
mammals. On the other hand, immunohistochemical staining (peroxidase anti-peroxidase, PAP, technique) showed localization of immunoreactive TRH (IR-TRH)
within specific regions of the hypothalamus, median eminence, and other areas of
the amphibian brain, thus providing an
anatomical basis for involvement of TRH
in metamorphosis (Mimnagh et al., 1987).
So, the question remains: does TRH serve
a neuroendocrine role in regulation of
metamorphosis?
Since concentrations of IR-TRH are
much higher in frog skin than brain (review:
Jackson and Bolaffi, 1983), we also deter-
771
mined sites of IR-TRH within tadpole and
frog skin, using two different techniques,
PAP and immunogold with silver enhancement. Both procedures demonstrated that
serous (granular) glands are the main sites
of TRH in amphibian skin (Fig. 10). Moreover, IR-TRH is present in such glands as
soon as they develop in the tadpole (Stroz
and Kaltenbach, 1986). Immunostaining
has provided an example of a single peptide (TRH) present both in skin glands and
in the nervous system. Is TRH also localized in digestive glands of tadpoles or frogs?
What function may it play in Amphibia?
Questions to be answered are endless.
In conclusion, I hope that the present
discussion of recent major advances in hormone research has heightened your awareness of the complexities of endocrine regulation of the internal environment in
vertebrates.
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