Physiol Rev
82: 875– 891, 2002; 10.1152/physrev.00010.2002.
Transport-Mediated Synapses in the Retina
E. A. SCHWARTZ
Department of Neurobiology, Pharmacology, and Physiology, University of Chicago, Chicago, Illinois
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Schwartz, E. A. Transport-Mediated Synapses in the Retina. Physiol Rev 82: 875– 891, 2002; 10.1152/physrev.00010.2002.—Most synapses rely on regulated exocytosis for determining the concentration of transmitter in the
synaptic cleft. However, this mechanism may not be universal. Several synapses in the retina appear to use a
synaptic machinery in which transmitter transporters play an essential role. Two types of transport-mediated
synapses have been proposed. These synapses have been best observed in horizontal cells and cones of nonmammalian retinas. Horizontal cells use a transporter to mediate a bidirectional shuttle, whose balance point is set by ion
concentrations and voltage. Nonmammalian cones combine exocytosis and the activity of a transporter. Because
exocytosis is voltage independent over most of a cone’s physiological voltage range, a voltage-dependent transporter
determines the concentration of transmitter in the synaptic cleft. These two synapses may be models for transportmediated synapses that operate in other parts of the brain.
I. INTRODUCTION
Most neuroscientists share a common mental picture
of the specialized region where information passes rapidly from one neuron to another. Elegant experiments
first defined the basic properties of chemical transmission
(66). Subsequently, the mental picture has been refined by
a wealth of anatomical, electrical, biochemical, and molecular biological experiments. The idealized synapse is a
fine model for the behavior of most real synapses. Consequently, it is often regarded as the only mechanism for
communication between neurons. But a few synapses
have been identified where the standard model appears to
fail. Several of these use a synaptic machinery in which
transmitter transporters play an essential role. The best
examples have been found in the retina. This may not be
an accident. First, the anatomy and physiology of the
retina have been intensively studied and are better known
than, perhaps, any other part of the brain. Second, cells in
the outer retina communicate without action potentials
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and, instead (as described in Fig. 4 and sect. II, A6 and
B1), control the release of transmitter with both hyperpolarizing and depolarizing changes in voltage that last
tens to hundreds of milliseconds. As I shall explain, transport-mediated synapses work best when controlled by
slow changes in presynaptic voltage. Two types of transport-mediated synapse have been proposed. Here, I first
describe the properties of the standard synapse and then
the idealized properties of the transport-mediated synapses. Afterwards, I consider the evidence. The discussion includes the twists and detours necessary to understand both mechanism and function.
A. Standard Synapse
The standard synapse relies on regulated exocytosis
(Fig. 1A). Vesicles, 30 – 60 nm in diameter, are first filled
with transmitter molecules and docked at specialized “active zones” where they wait for a signal. The release of a
vesicle’s contents commences when depolarization opens
0031-9333/02 $15.00 Copyright © 2002 the American Physiological Society
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I. Introduction
A. Standard synapse
B. What are transporters?
C. Transport shuttle
D. Transport retrieval
E. Comparison
II. Examples of Transport-Mediated Synapses
A. Transport shuttle in horizontal cells
B. Transport retrieval in cones
C. Other synapses?
III. Questions and Conclusions
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E. A. SCHWARTZ
Ca2⫹ channels and Ca2⫹ flows into the cytoplasm. Ca2⫹ is
the immediate trigger for the fusion of vesicle and surface
membranes. Transmitter molecules exposed to the extracellular space quickly diffuse to receptors on nearby neurons. The entire release process can be completed in ⬍1
ms (52, 80). Subsequently, transmission is terminated as
transmitter molecules diffuse from a release site, are
chemically modified, or removed from the extracellular
space by transporters. Although transporters help remove
transmitter molecules from the synaptic space, their role
is not essential for the rapid transfer of synaptic information.
Three properties are characteristic of the standard
model. First, exocytosis is triggered by Ca2⫹ influx and a
rise in intracellular Ca2⫹ concentration (37, 67). Second,
each loaded vesicle contains a few thousand transmitter
molecules (69). As a consequence, the concentration in
the synaptic cleft can momentarily reach a relatively high
concentration, i.e., ⬎1 mM (24). Third, because all loaded
vesicles contain a roughly equal number of transmitter
molecules, sequential fusion events are of roughly equal
size (31), i.e., release is “quantized.”
B. What Are Transporters?
Charged molecules, like transmitters, do not easily
penetrate lipid membranes. Instead, channels, pumps,
and transporters provide substrate-specific pathways
through a membrane. Channels are aqueous pores that
allow ions to move down an electrochemical gradient.
Pumps utilize energy derived from the breaking of chemical bonds to do the work required to move a substrate up
a concentration gradient. Transporters can couple the
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movements of several substrates. The entire process can
be considered to be a chemical reaction that translocates
reactants, transmitter and ions, across a membrane rather
than altering chemical bonds. Some reactants will move
down their electrochemical gradient while others are
moved up, so long as the total movement is energetically
favorable. The transport reaction is reversible. Equilibrium occurs when the forward and backward fluxes are
equal. If the reactants are charged, zero current is observed at a reversal potential. Away from the reversal
potential, the rate of transport is determined both by
membrane voltage and the concentrations of all of the
reactants.
Transporters can be grouped into gene families
whose members share sequence similarity. Na⫹-coupled
transporters are a large family whose members move
several transmitters across the surface membrane (96). In
humans there are four GABA transporters, two glycine
transporters, one for monoamines, and another for serotonin. A separate family of five transporters translocates
glutamate (128). In addition, H⫹-coupled transporters are
preferentially expressed in synaptic vesicles (41, 45).
Transporters have distinctive ion requirements. Each
Na⫹-coupled transporter moves a substrate (transmitter)
with Na⫹ and a select set of other ions across a cell’s
surface membrane (see Ref. 117). The requirement for
Na⫹ is stringent. No other ion can substitute. Consequently, the absolute dependence on Na⫹ can be used as
a diagnostic test.
Each transport reaction moves only one or a small
number of transmitter molecules across a membrane. The
maximum reaction rate has been estimated as 10 –100 s⫺1.
This estimate often comes from the time course of a
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FIG. 1. Model synapses. Bold arrows indicate voltage-dependent events. A: the standard synapse uses exocytosis.
Depolarization opens channels that allow the entry of Ca2⫹ (open circles) into the cytoplasm. An increase the Ca2⫹
concentration near release sites triggers the exocytosis of vesicles containing transmitter molecules (solid circles). B: a
transport shuttle uses a voltage-dependent transporter (cogged wheel) to move transmitter in both directions across the
cell membrane. Hyperpolarization speeds uptake, and depolarization speeds release. C: transport retrieval combines a
steady, high rate of voltage-independent exocytosis with a voltage-dependent uptake. A voltage-independent channel
continuously allows Ca2⫹ to enter the cytoplasm. A voltage-dependent transporter removes transmitter molecules from
the synaptic cleft. The balance between a steady exocytosis and a variable rate of uptake determines the concentration
of transmitter in the cleft.
TRANSPORT-MEDIATED SYNAPSES IN THE RETINA
C. Transport Shuttle
The model transport-synapse incorporates a bidirectional shuttle whose balance point is set by ion concentrations and voltage (Fig. 1B). Intra- and extracellular
transmitter concentrations are in dynamic balance. Usually the intracellular concentration is high, perhaps 10
mM, while the extracellular concentration is low, ⬍100
␮M. In addition, the intracellular volume is relatively large
compared with the much smaller extracellular volume. A
low concentration in a small extracellular volume is buffered by a high concentration in a large intracellular volume. Altering the balance between influx and efflux
changes the concentration of transmitter in the extracellular space.
Two properties are characteristic of a transport shuttle. First, Na⫹ is required for both substrate influx (uptake) and efflux (release). Release occurs only when a
neuron has adequate transmitter and Na⫹ concentrations.
Second, transporters are less sensitive than voltage-gated
channels to changes in membrane voltage. Consequently,
a transport shuttle is likely to require a relatively large
change in either presynaptic voltage or ion gradients.
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Transport-mediated release lacks the anatomical specialization of vesicles localized at a release site. Thus the
identification of a transport synapse usually depends on
physiology. Three observations can help in the identification. 1) Transport requires Na⫹ and does not depend on
Ca2⫹. 2) The voltage dependence for transport does not
match the voltage dependence of the Ca2⫹ current. 3)
Transporters have a distinctive pharmacology. Selective
and potent agents are available for catecholamine and
indoleamine transporters (14). A smaller pharmacopoeia
is available for GABA transporters (16). Unfortunately,
equally good drugs are not yet available for glutamate
transporters (30).
D. Transport Retrieval
A second type of transport synapse is a hybrid that
depends on a balance between exocytosis and transport.
Although exocytosis releases transmitter, it is voltagedependent uptake that is essential for controlling the
concentration of transmitter in the synaptic cleft. In an
extreme case, a steady rate of vesicle fusion continuously
adds transmitter to the synaptic cleft. The concentration
of transmitter in the cleft is determined only by the rate of
transmitter removal (Fig. 1C). Hyperpolarization speeds
the transporter and lowers the concentration in the cleft,
and depolarization has the opposite affect.
Standard and transport-retrieval synapses use exocytosis in different ways. The standard synapse relies upon
voltage-gated Ca2⫹ entry to produce a rapid fluctuation in
intraterminal Ca2⫹ concentration and a burst of exocytosis. The transport-retrieval synapse maintains a relatively
constant intraterminal Ca2⫹ concentration and produces
a steady stream of vesicle fusion. Continuous exocytosis
is unlikely at a synapse with only a few fusion sites but
can occur in retinal neurons with ribbon synapses (see
below), which have hundreds or even thousands of vesicle fusion sites (112, 151).
Transport-retrieval requires the following: 1) a surfeit
of vesicles and hundreds of fusion sites able to sustain
high rates of continuous exocytosis and 2) a voltagedependent transporter in the presynaptic membrane that
can control the transmitter concentration in the synaptic
cleft.
Transporters play different roles in the transportshuttle and transport-retrieval synapses. A transport shuttle moves transmitter in both directions across the presynaptic membrane. The transporter at a transportretrieval synapse needs to operate in only one direction,
removing transmitter from the synaptic cleft. Accordingly, reverse transport is unnecessary, and the intracellular transmitter and/or Na⫹ concentrations can be relatively low.
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transient current observed in the absence of transmitter
(82, 152). However, a reaction observed in the absence of
transmitter may not indicate the rate of transport when
transmitter is present. In fact, transporters may operate at
rather brisk rates. For example, a Ca2⫹ transporter operates at rates of 5,000 s⫺1 (53). Likewise, a GABA transporter can cycle at a rate of ⬃1,000 s⫺1 (unpublished
observations; see also Ref. 4 for evidence that the partial
reaction for influx through a glutamate transporter operates at ⬃1,000 s⫺1).
Transporters were initially viewed as molecular machines that moved substrate from the extracellular medium into the cytoplasm. “Reverse” transport, viz. efflux,
was rarely encountered and considered to be inefficient.
Why? Efflux is usually limited by the intracellular concentrations of Na⫹ and transmitter. The consequence can be
emphasized with two examples. Transporters allow Müller glia to remove glutamate from the retina’s extracellular space. The efficient intracellular conversion of glutamate to glutamine keeps the cytoplasmic glutamate
concentration ⬍0.1 mM (84, 85). Moreover, the intracellular cation is predominately K⫹ rather than Na⫹. Low
concentrations of intracellular glutamate and Na⫹ make
efflux unlikely during physiological conditions. Next consider a retinal horizontal cell that accumulates GABA. The
intracellular GABA concentration may be 10 mM (72, 84).
Moreover, Na⫹ enters continuously through synaptic receptors activated by the release of glutamate from adjacent photoreceptors. Thus a horizontal cell contains both
GABA and Na⫹. As a result, the efflux of GABA from
horizontal cells is possible.
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parts deserve a careful review to see what is known, what
is speculated, and what is disputed.
E. Comparison
II. EXAMPLES OF TRANSPORT-MEDIATED
SYNAPSES
A. Transport Shuttle in Horizontal Cells
The synapse between horizontal cells and cones provides, perhaps, the best-studied case for transport-mediated release. The story might at first seem simple. Horizontal cells accumulate and release GABA. Postsynaptic
cones have GABA receptors. The story would be humdrum except for the claim that transmitter release is
transport mediated. This unusual proposal requires an
extraordinary proof. Although many parts have been confirmed and indirect evidence is abundant, a rigorous demonstration that transporters mediate synaptic communication has not been accomplished. Therefore, all of the
1
Fast synaptic transmission uses AMPA receptors, with an EC50
between 80 and 300 ␮M glutamate. NMDA and metabotropic receptors
have an EC50 between 1 and 4 ␮M (35, 99, 121).
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1. An atypical synapse
Synaptic interactions between cones, horizontal, and
bipolar cells create the first step in processing visual
information. Horizontal cells form an electrical syncytium, which integrates input from photoreceptors over a
large retinal area. This integrated activity is relayed back
to cones and forward to bipolar cells as a “surround
signal.” Horizontal cells and bipolar cells poke slender
fingers into a cone’s synaptic ending. Each photoreceptor
has 5–25 invaginations. The typical organization within
each invagination is a triad (89, 131), a central bipolar cell
process sandwiched between two lateral, horizontal cell
processes (Figs. 2 and 3). The synaptic site is clearly
marked by a cytoplasmic organelle, a proteinaceous sheet
or “ribbon” that is anchored to the presynaptic membrane. Vesicles fill a cone’s cytoplasm and are tethered to
the flanks of ribbons. Several rows of vesicle fusion sites
dimple the surface membrane along the flanks of a ribbon
(112). In contrast, the lateral (horizontal cell) process is
either empty (Fig. 3B) or occasionally contains scattered
vesicles (Fig. 3C). An active zone and fusion sites are
absent from the horizontal cell’s surface membrane (112).
In many species these empty processes are the only sites
where cones and horizontal cells are sufficiently close to
form a synapse.
The lateral processes of horizontal cells (Fig. 3) provide an exception to the nearly monotonous morphology
of conventional synapses. The absence of both vesicle
clusters and fusion sites has been a lump under the carpet
on which discussions of synaptic mechanism have tripped
for nearly 20 years. At first the anatomy was only puzzling.
The notion that these were sites of a novel release mechanism began with the observation that the putative transmitter GABA could be released without Ca2⫹. A sheet of
horizontal cells was isolated from the toad retina by killing other retinal neurons with a cocktail of toxins (124).
The surviving horizontal cells released GABA when depolarized either by K⫹ or glutamate. The release of GABA
was unchanged when Ca2⫹ was removed from the extracellular saline and replaced with Co2⫹, an ion that blocks
voltage-dependent Ca2⫹ channels and does not support
exocytosis.
The argument for transport-mediated release depends on the following observations: 1) GABA is synthesized and accumulated in horizontal cells, 2) GABA is
released from horizontal cells during darkness, 3) the
release of GABA is Na⫹ dependent and Ca2⫹ independent,
4) horizontal cells express a GABA transporter that mediates reverse transport in both intact retinas and isolated
cells, and 5) GABA receptors are expressed at postsynaptic sites. Results supporting these points have come
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Regulated exocytosis is the explosive, one-way delivery of a few thousand molecules into the synaptic space.
It is ideally suited to transmit the staccato information
encoded by action potentials. The pulsatile mobilization
of quanta produces brief and high transmitter concentrations (24). Repeated stimulation often produces a series
of unequal responses that reflect a random fluctuation in
the number of released quanta (see Ref. 5).
Transporters operate in a different quantitative regime. Each transport reaction moves only one or a few
transmitter molecules across a membrane. The total flux
depends on the density of transporters and the rate of
translocation. Even at a high density, a transporter with a
slow cycle time may not be able to make rapid changes in
extracellular transmitter concentrations. Hence, transport-mediated release is best controlled by relatively slow
changes in voltage or ion concentrations. Consequently, a
transport synapse is more likely to be found when
changes in presynaptic voltage persist for hundreds of
milliseconds rather than 1–10 ms. Slow, small undulations
in extracellular transmitter concentration are in stark
contrast to the large, brief injections produced by exocytosis. Modest changes in transmitter concentrations must
be coupled to postsynaptic receptors with a relatively
high sensitivity and little desensitization. Postsynaptic receptors for GABA, catecholamines, and glycine (or Dserine) are likely targets. On the other hand, the lower
affinity and rapid desensitization of ionotropic glutamate
receptors makes them unlikely targets of a transport synapse (however, see sect. IIB1).1
TRANSPORT-MEDIATED SYNAPSES IN THE RETINA
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from many species. One problem is to separate consistent
features seen in most retinas from variations and specializations that distinguish individual species. I first discuss
results from four nonmammalian species. No single species provides a complete set of results. Instead, their
contributions articulate like pieces of a jigsaw puzzle.
2. Turtle
Two hundred years ago Thomas Young (170) assumed that the response in a photoreceptor was determined only by the light it absorbed, an idea later incorporated into the Young-Helmholtz theory of trichromatic
vision. Consequently, it was a surprise when Baylor et al.
(12) reported that cones also receive a synaptic input
from horizontal cells. The processing of visual information begins when each cone combines the hyperpolarization produced by the light it absorbs with a synaptic input
from horizontal cells, which relays information about the
average light intensity that falls on a large area of surrounding retina. The “surround signal” was first seen in
the turtle retina.
The turtle has four types of horizontal cell (see Ref.
28). Only one type uses a transporter to accumulate
[3H]GABA (71). The same cells also express an enzyme
required for GABA synthesis: glutamic acid decarboxylase (GAD) (55). GABA, unlike glutamate or glycine, is not
an amino acid involved in intermediary metabolism. It is
synthesized only in neurons and ␤-cells of the pancreas,
where its presence usually indicates a role in intercellular
communication.
Physiology almost implicates GABA in the synaptic
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communication between horizontal cells and cones. Exogenous GABA depolarizes horizontal cells and attenuates and slows responses elicited by flashes of light (104).
Isolated cones, but not rods, have GABA receptors on
their synaptic endings (64). These receptors were believed to be sites of synaptic input. However, inexplicably,
GABA agonists and antagonists have been reported to
have no effect on the synaptic signal transmitted from
horizontal cells to cones (139). It is hard to understand
how GABA agonists can alter the amplitude and time
course of response in a presynaptic horizontal cell (104),
and increase the conductance of a postsynaptic cone (64),
but not alter the synaptic signal that passes from the
horizontal cell to the cone (139). Because the results are
inconsistent, they almost, but not quite, implicate GABA
as a transmitter. Fortunately, results from the salamander
help fill the gap.
3. Salamander
The salamander retina has two types of horizontal
cells. One type expresses GAD (159) and accumulates
[3H]GABA (155). The presence of a GABA transporter is
confirmed by the ability of a relatively specific inhibitor,
NO-711, to block a GABA-activated current (161; but see
Ref. 81). Transport and synthesis produce a high cytoplasmic GABA concentration (32, 77, 155, 159).
The combination of immunohistochemistry, physiology, and pharmacology has demonstrated that GABA mediates signals transmitted to cones, bipolar cells, and
horizontal cells. GABA receptors decorate photoreceptor
terminals and horizontal cell processes (153). The sur-
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FIG. 2. Cartoon to explain synaptic communication at
the cone’s invaginating synapse. C, cone; H, horizontal cell;
B, bipolar cell. A synaptic ribbon in the cone cytoplasm
marks the site of vesicle release. Three processes from
postsynaptic cells are arranged in an indentation (or invagination) opposite a ribbon. The central process is a bipolar
cell dendrite. The two lateral processes are from horizontal
cells. Horizontal cells receive an excitatory (sign-preserving) signal from cones and return an inhibitory (sign-reversing) signal to cones. The result is a “feedback” pathway. There are two types of bipolar cell: “on” bipolar cells
[B(on)] are inhibited by cones and excited by horizontal
cells, and “off” bipolar cells [B(off)] receive the opposite
polarity signals. Direct input from horizontal cells to bipolar cells creates a “feed-forward” pathway. An individual
cone makes contact with both types of bipolar cell.
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round signal transmitted to cones is suppressed by
GABAA agonists and antagonists (156). In a similar manner, surround signals transmitted to bipolar cells are inhibited by GABA (155) or a combination of GABAA and
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GABAB antagonists (49). In addition, physiology demonstrates autoreceptors on isolated horizontal cells (47),
which shape responses to flashes of light in intact retinas
(63). When GABA is added to saline superfusing an iso-
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FIG. 3. A: low-power electron micrograph of a cone’s synaptic pedicle in the
turtle (Pseudemys scripta elegans) retina. An arrow points to a ribbon with a
halo of synaptic vesicles. Three ribbons
are evident. The cone’s cytoplasm is
filled with synaptic vesicles. B: higher
power view of the invaginating complex
in the turtle retina. C: the invaginating
complex in the monkey (Macaca arctoides) retina. H, horizontal cell lateral
process; B, bipolar cell dendrite. The lateral processes made by horizontal cells
are usually empty (as in B) or occasionally contain scattered vesicles (as in C).
There are no obvious active zones in the
lateral processes. (Electron micrographs
kindly provided by Elio Raviola, Harvard
Medical School.)
TRANSPORT-MEDIATED SYNAPSES IN THE RETINA
881
lated retina, responses produced by flashes of light are
attenuated and slowed (63, 162, 163). Because similar
changes occur during dark adaptation, Yang and Wu (160)
suggest that dark adaptation increases extracellular
GABA concentration.
Although several details must still be resolved, the
experiments described above indicate that GABA is actually used as a horizontal cell transmitter. Nonetheless, the
salamander has provided little insight into the mechanism
of GABA release. This part of the story has come from
other species, particularly the catfish.
4. Catfish
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FIG. 4. Membrane currents that directly influence synaptic communication during physiological function: comparison of voltage responses
produced by light and membrane currents. The results are from two
different types of preparations: 1) voltage responses (black traces)
produced by a flash of light (indicated by the purple timing traces) were
recorded after cells in an eyecup were impaled with a sharp pipette.
Voltage is plotted along the abscissas against time as the left ordinate. 2)
Membrane currents (colored traces), measured in solitary cells during
whole cell recording, are plotted along the right ordinate. A: data from
catfish (Ictalarus punctatus) horizontal cells. The response to light
(black trace) begins at a resting potential of ⫺21 mV and reaches a peak
voltage of ⫺75 mV. The red trace is the voltage-gated Ca2⫹ current (ICa).
The green trace is the efflux current produced by the GABA transporter
when the intracellular solution contains GABA. The blue trace is the
influx current produced by extracellular GABA. B: data from salamander
(Ambystoma tigrinum) cones. The response to light (black trace) begins at a resting potential of ⫺39 mV and reaches a peak voltage of ⫺62
mV. The red trace is ICa. The green trace is the cGMP-gated current.
Currents are normalized to their maximum amplitude. The cGMP current is usually ⬍5 pA; the maximum amplitude of ICa is often larger than
100 pA. For comparison, the blue trace is ICa recorded from a cone in the
squirrel (Spermophilus tridecimlineatus) retina.
GABA produces an outward current at depolarized potentials (22). Similar currents have been observed when the
rat GAT-1 transporter was expressed in cultured cells (21,
116). The transporter shuttles GABA in both directions
across the plasma membrane (see Fig. 4A for the voltage
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The catfish retina has two types of horizontal cells.
Rods selectively contact one type of horizontal cell, and
cones contact the other type. Only the cone-driven horizontal cells accumulate GABA (72). The cytoplasmic concentration of GABA is estimated to be ⬃10 mM (73).
Experiments with intact retinas provide a broad outline of
GABA movements (73, 75). The intracellular concentration is highest during constant illumination, when horizontal cells are hyperpolarized, and decreases during
darkness, when they are depolarized. Presumably GABA
is transferred from the cytoplasm to the extracellular
space, where it is used as a neurotransmitter. As expected, surround signals in cone photoreceptors are
blocked by a GABA receptor antagonist (72, 75).
Information about the mechanism of release comes
from isolated cells. The release of GABA can be measured
with a sensitive detector. Goldfish bipolar cells have a
large GABAA current. When a catfish horizontal cell and a
goldfish bipolar cell were isolated, placed in the same
dish, and then pushed against each other, the activation of
a current in the bipolar cell could be used to detect the
release of GABA from the horizontal cell (127). Even after
Ca2⫹ in both the external solution and the horizontal cell’s
internal solution was buffered to 50 nM with a combination of EGTA and BAPTA, GABA was released when the
horizontal cell was depolarized. Thus depolarization released GABA by a mechanism that did not require a
change in intracellular Ca2⫹ concentration. Moreover, the
release of GABA increased as the horizontal cell was
depolarized from ⫺80 to ⫹20 mV and did not correlate
with the voltage dependence of the Ca2⫹ current (Fig. 4A,
red trace). The release of GABA has since been found to
closely match the voltage dependence of efflux through
the GABA transporter (Fig. 4A, green trace; compare with
Fig. 3B in Ref. 127).
Isolated horizontal cells have two GABA-activated
currents (22, 36, 60, 136, 137). A large component is
produced by GABA-activated channels, which act as autoreceptors. A smaller component is produced by an electrogenic transporter. Extracellular GABA produces an inward current at hyperpolarized potentials, and intracellular
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5. Goldfish
The model transport-mediated synapse incorporates
the release mechanism observed in isolated, catfish horizontal cells and the surround signal mediated by GABA in
salamander retinas. Experiments with goldfish retinas
echo results from catfish and salamanders.
The goldfish has four morphological types of horizontal cells. One of these receives input from rods, and three
types receive input from varying combinations of cones
(132, 133). Only one type of cone-driven horizontal cell
expresses GAD (10, 17) and accumulates GABA (84, 86,
94, 101, 169). GABA is accumulated in the cytoplasm
during continuous illumination, when horizontal cells are
hyperpolarized, and is released during darkness, when
they are depolarized (86, 165, 168). In addition, release is
Ca2⫹ independent and increased when horizontal cells
are preloaded with Na⫹ (164, 166). Experiments on isolated cells have demonstrated that a transporter is responsible for the efflux of GABA. Retinas were dissociated,
and horizontal cells were separated from other cells by
velocity sedimentation (6 – 8). GABA was released when a
collection of several hundred isolated cells was depolarized with a high-K⫹ saline or glutamate. Most of the GABA
efflux was Na⫹ dependent, Ca2⫹ independent, and controlled by voltage over a broad range. Only a small part
was Ca2⫹ dependent. Voltage-dependent transport is the
predominant mechanism controlling the movement of
GABA between cytoplasm and the extracellular space.
Physiology and pharmacology experiments, similar
to those already described for the salamander and catfish,
indicate that GABA is used as a horizontal cell transmitter
and contributes to the surround signal. GABA affects the
responses to light of both cones and horizontal cells
(157). In a related teleost, the carp, GABA mediates a
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surround signal transmitted from horizontal cells to cones
(95, 142, 143).
6. Role of GABA in nonmammalian horizontal cells
All of the items necessary for transport-mediated
release are present in nonmammalian horizontal cells. A
phylogenetic survey of vertebrates demonstrates that
GABA is synthesized and accumulated in cone-driven horizontal cells in virtually all nonmammalian retinas (83).
This includes cartilaginous and bony fish, amphibians,
reptiles, and birds. The intracellular concentration of
GABA depends on membrane voltage. Light produces a
change in membrane potential that is well suited for
controlling a voltage-dependent transporter. Like other
neurons in the outer retina, horizontal cells do not produce action potentials, but instead generate potentials
that are continuously graded between ⫺20 and ⫺70 mV.
Shifts in voltage occur on a time scale of ⬃100 ms. Large,
graded potentials are ideal for controlling a transport
mechanism. GABA is released when horizontal cells are
depolarized. The voltage dependence of Na⫹-coupled
GABA transporters is sufficient to control the flux of
GABA. Once released, GABA activates receptors on cones
and bipolar cells as well as autoreceptors on horizontal
cells.
The first tendency was to believe that GABA was the
whole story and able by itself to explain the genesis of
surround signals and all lateral interactions in the outer
retina. The following considerations, all based on observations made in the goldfish retina, provide a slightly
different view.
Synaptic interaction between a horizontal cell and a
cone was initially assumed to occur within the invagination. However, ultrastructural immunolabeling demonstrates few GABAA receptors located within the invaginating complex. Instead, they are located on the lateral
surfaces of the cone’s synaptic pedicle (169). GABA must
diffuse in the extracellular space to reach these receptors.
The release of GABA from the entire cell surface (see
sect. IIA4) together with GABA receptors at a “peri-synaptic” location implies that GABA operates by volume
transmission (for a discussion of volume transmission,
see Ref. 43).
There is a gradient in GABAergic properties between
central and peripheral retina. Horizontal cells in the periphery accumulate more [3H]GABA and label more intensely with GABA and GAD antibodies than centrally
located cells (S. Yazulla, personal communication; see
also Ref. 1). The function of horizontal cells in the retinal
circuit may change with retinal position.
The activity of the horizontal cell’s GABA transporter
may be modulated with a circadian rhythm. In teleosts,
dopaminergic interplexiform cells cross from the inner to
the outer plexiform layer and make abundant synapses on
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dependence of the influx and efflux currents). The net flux
is controlled by membrane voltage.
Experiments on isolated catfish horizontal cells indicate that GABA transporters are expressed at a reasonably high density over a large part of the surface membrane (22, 36, 60, 136, 137). Consequently, transporters
allow a depolarized horizontal cell to ooze transmitter
from its entire surface. Although this is not a picture of
selective communication restricted to specialized points
of contact, the cloud of transmitter that surrounds a
horizontal cell may be what is required to transfer information about the horizontal cell membrane potential to
neighboring cells.
Catfish horizontal cells have provided an important
piece of the jigsaw puzzle. The concentration of GABA in
horizontal cells changes with illumination. A Na⫹-coupled
transporter moves GABA in both directions across the
membrane. The amplitude and direction of net flux is
controlled by membrane voltage.
TRANSPORT-MEDIATED SYNAPSES IN THE RETINA
7. Mammalian horizontal cells
Nonmammals have between two and five types of
horizontal cells. Usually, only one type of cone-driven cell
is GABAergic. In contrast, most mammals have two types
of horizontal cells, and both are GABAergic. Our understanding of the role of GABA in mammalian horizontal
cells has changed during the past 10 years. Mammalian
horizontal cells usually contain less GABA than GABAergic amacrine cells in the same retina. In addition, horizontal cells in different species have very different apparent GABA concentrations. The rabbit and cat illustrate
this situation. The rabbit typifies mammalian species in
which GABA content is relatively low. It is now apparent
that the concentration of GABA is developmentally regulated. At first, during an early phase of retinal differentiation, both types of horizontal cell accumulate [3H]GABA
(113) and contain an appreciable GABA concentration
(88, 101, 109). Afterwards, uptake and GABA content
decline as Müller glia differentiate and become new sites
of GABA uptake. Although adult horizontal cells continue
to express the synthetic enzyme GAD (59, 93), the concentration of GABA in adult horizontal cells is relatively
low (59), albeit greater than in non-GABAergic neighbors.
The cat typifies a mammal whose horizontal cells contain
a significant concentration of GABA. The expression of
GAD has been demonstrated by immunolabeling (145)
and by in situ mRNA hybridization (119). The concentration of GABA is moderate but still less than in amacrine
cells (85, 107, 145, 154).
Although their intracellular GABA concentration appears to be low or modest, there is reasonable evidence
that mammalian horizontal cells use GABA as a neurotransmitter. For example, immunohistochemistry labels
GABAA receptors on rabbit cone synaptic terminals (90)
and the shafts of bipolar cell dendrites adjacent to cone
pedicles in the rabbit (48) and cat (146) retinas. Complimentary evidence comes from physiology. GABA activates autoreceptors in isolated, rabbit horizontal cells
(15), and both attenuate and slow responses produced in
Physiol Rev • VOL
intact retinas (15). Likewise, GABAA agonists have a similar effect on cat horizontal cells (42). Thus there is anatomical and physiological evidence for GABA as a horizontal cell transmitter in both nonmammalian and
mammalian retinas.
8. A difference between nonmammals and mammals
There is a significant difference in the chemical organization of nonmammalian and mammalian retinas that
involves both horizontal cells and Müller glia (see Ref.
83). Müller glia in nonmammalian retinas do not take up
GABA. At the same time, virtually all nonmammals have
one type of horizontal cell that synthesizes and accumulates a high concentration of GABA. The situation is altered in mammals, which have Müller glia that avidly
scavenge GABA from the extracellular space and horizontal cells with only a low to moderate GABA concentration.
As expected, a Na⫹-coupled transporter moves
GABA into mammalian Müller glia. Unexpectedly, there is
no indication of a Na⫹-coupled transporter in mammalian
horizontal cells. Johnson et al. (58) labeled rat retinas
with specific antibodies that recognize three GABA transporters. Müller glia were labeled by two of these antibodies, and horizontal cells were unlabeled. A novel activitydependent assay provided a similar result (108). The
uptake of GABA transport agonists (␥-vinyl GABA, diaminobutyric acid, and gabaculine) was detected with specific antibodies. Amacrine and Müller cells were labeled
in cat and monkey retinas, but horizontal cells remained
unlabeled.
Nonmammalian horizontal cells control the GABA
concentration in synaptic and nonsynaptic spaces. In
mammals, these tasks appear to be divided between horizontal cells and Müller glia. Mammalian Müller glia use a
Na⫹-coupled transporter to control the extracellular
GABA concentration in nonsynaptic regions. Mammalian
horizontal cells may use another mechanism to focally
release GABA at synaptic sites. Monkey horizontal cells
express an H⫹-coupled “vesicular” GABA transporter,
VGAT. Light-microscope immunohistochemistry reveals a
low density expressed in the cell surface and a higher
density in dendrites that poke into the invaginating synapse (51). A similar distribution of VGAT is also seen in
the rat, mouse, and rabbit retinas (29). Intense labeling of
dendrites may reflect a high density in the surface membrane or an aggregation of synaptic vesicles. Electronmicroscope immunohistochemistry will be needed to determine the site. Because the voltage and ion dependence
of the H⫹-coupled transporter are incompletely known, it
is not yet clear whether this transporter could mediate the
voltage-dependent release of GABA.
Both nonmammalian and mammalian horizontal cells
appear to use GABA as a transmitter. A Na⫹-coupled
transporter releases GABA from the entire cell surface of
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cone-driven horizontal cells (141). The release of dopamine follows a circadian cycle (see Ref. 19). Exogenous
dopamine inhibits the release of GABA from horizontal
cells (167; see also Ref. 65). As a consequence, the release
of GABA may be reduced during the day and increased at
night.
These considerations suggest that GABA communication involves volume transmission, is accentuated in
the periphery, and is modulated in a circadian cycle.
Although GABA may still contribute to a surround signal,
this more restricted picture of GABA action makes it
unlikely that the transport-mediated release of GABA can
explain all synaptic interactions mediated by horizontal
cells.
883
884
E. A. SCHWARTZ
one type of cone-driven horizontal cell in nonmammalian
retinas. Although mammalian horizontal cells appear to
use GABA, another mechanism may restrict release to the
tips of their dendrites.
role of GABAergic transmission may require understanding the entire circuit.
9. Conundrums
Our perspective of GABA and transport-mediated release by horizontal cells will be altered if we discover
other mechanisms that contribute to the surround signal.
Hence, I include a detour to describe an ephaptic mechanism that has recently been suggested (61). No transmitter is involved. A horizontal cell dendrite that pokes into
a cone’s synaptic base (see Figs. 2 and 3) is assumed to
create an extracellular path with a high electrical resistance. The current that passes out the tip of a dendrite
produces a voltage as it traverses the extracellular resistance. For example, 100 pA flowing out of a horizontal
dendrite and into an extracellular space with 1-M⍀ resistance would produce an extracellular voltage of 0.1 mV.
As a consequence, channels in the cone membrane that
are located opposite the current source would be depolarized by the shift in extracellular voltage, in this case
0.1 mV.
Ephaptic transmission within the cone invagination
was originally suggested by Byzov and Shura-Bura (20).
The most recent version, proposed by Kamermans et al.
(61), emphasizes a special role for hemi-gap channels. An
antibody that recognizes the gap-junction protein connexin-26 (Cx26) labels the tips of goldfish horizontal cell
dendrites but does not label gap junctions that connect
neighboring horizontal cells (57). Because electron microscopy fails to provide evidence for gap junctions
within the invagination, Kamermans et al. (61) suggest
that Cx26 forms hemi-gap channels. Normally, hemi-gap
channels are kept closed by a negative membrane potential and extracellular Ca2⫹ (33). Kamermans et al. (61)
suggest that Cx26 forms patent hemi-gap channels in
horizontal cell dendrites. They buttress their claim with a
physiological experiment. After retinas were continuously
treated with SKF89976A, a potent inhibitor of GABA
transport, a component of the surround signal survived.
Carbenoxolone, an agent that closes gap junction channels, blocked the surviving surround signal. The anatomical and physiological correlation strikes one as more
than a coincidence and is the essence of the argument for
ephaptic transmission.
Ephaptic transmission through hemi-gap channels
creates at least three problems.
1) A significant change in extracellular voltage requires either a very large current or an improbably large
extracellular resistance (see above). Moreover, it is difficult to see how hemi-gap channels can be the source of a
large extracellular current and glutamate receptors can
simultaneously polarize a cell from ⫺70 to ⫺20 mV without violating Ohm’s law.
2) Hemi-gap channels are permeable to molecules
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The exact part that transport-mediated release of
GABA plays in synaptic interactions between horizontal
cells and cones continues to be a vexing question. The
following observations indicate some of the difficulties
and problems that must be overcome to understand
GABA’s synaptic action.
Nonmammals have several subtypes of horizontal
cells. Each subtype may use different transmitters or
mechanisms for communicating with neighbors. Only one
type of cone-driven horizontal cell accumulates GABA.
For example, the goldfish has two types of cone-driven
horizontal cells and a rod-driven horizontal cell that do
not contain and release GABA. These cells must use other
transmitters or mechanisms for synaptic communication.
An individual horizontal cell may have more than one
mechanism for communicating with neighbors. For example, turtle (28), salamander (78), catfish (78), goldfish
(78), and rabbit (103) horizontal cells contain a nitric
oxide synthase. Presumably, these cells use nitric oxide to
alter the behavior of neighboring neurons.
Horizontal cells make their characteristic synaptic
connection in the invaginating complex. The typical lateral process has an empty cytoplasm (with few or no
vesicles) and lacks an active zone. In addition, in some
species, horizontal cells also make standard synapses at
other locations. For example, catfish horizontal cells
make conventional synapses (outside the invagination)
onto cone telodendria, fine dendrites that interconnect
neighboring cones (118). Because the catfish has only one
type of cone-driven horizontal cell, transport-mediated
and standard synapses must coexist in the same cell.
Another example is in the salamander retina, which has
two types of horizontal cells. One of these makes a conventional synapse with the second type (74). In this case,
it is not known which subtype accumulates GABA. Although these synapses may be peculiar to the species
cited, the examples illustrate that cone-driven horizontal
cells (or at least some of them) can make standard synapses. Finally, in the human retina, lateral processes that
contact rods (unlike those that contact cones) contain
vesicles and an active zone (79). The function of this
contact is unknown. Physiology has so far consistently
failed to demonstrate a surround signal in both nonmammalian and mammalian rods.
The transport-mediated release of GABA is only one
of several synaptic mechanisms that operate in the retina’s outer synaptic layer. Understanding the functional
10. An alternate mechanism for surround inhibition
885
TRANSPORT-MEDIATED SYNAPSES IN THE RETINA
11. Summary of transport-mediated release
by horizontal cells
One type of horizontal cell synthesizes, accumulates,
and releases GABA in virtually all nonmammals. GABA is
released during darkness when horizontal cells are depolarized and accumulated during illumination when they
are hyperpolarized. A voltage-dependent transporter operates for both influx and efflux. The transporter can
explain the release of GABA. However, the story is not
complete. The problem that remains is to identify exactly
what information is transmitted by the release of GABA.
The existence of several horizontal cell types with multiple synaptic interactions has complicated the analysis.
GABA transport is only expected in one subtype of horizontal cell. GABA has been reported both to be (49, 95,
142, 143, 155–157) and not to be (62, 81, 139) responsible
for a component of the surround signal. A rigorous demonstration that transporters mediate synaptic communication will require identifying synaptic signals that directly result from transport action. This will require
dissecting a GABA synaptic pathway from other mechanisms.
Although GABA is likely to be a horizontal cell transmitter in mammalian retinas, the role of transport-mediated release is less certain. Although the anatomy of the
mammalian retina has been intensively studied, its physiology has been, unfortunately, less well explored. Nonetheless, it is already apparent that there are significant
differences between nonmammalian and mammalian retinas.
Physiol Rev • VOL
B. Transport Retrieval in Cones
Cone photoreceptors provide another perspective on
transport-mediated synaptic transmission. The machinery
for exocytosis is obvious. The cytoplasm of a cone’s
synaptic ending is filled with synaptic vesicles, and intracellular ribbons mark active zones (Fig. 3A). Nonetheless,
two physiological observations in nonmammals suggest a
problem. First, the voltage dependence of the cone’s Ca2⫹
current (see Fig. 4B and Table 1) is barely activated
within the physiological operating range (76, 126). Second, a component of synaptic transmission is reported to
persist after Ca2⫹ entry through voltage-gated channels is
blocked with either Co2⫹ (125) or nisoldipine (115). These
observations can be resolved by the discovery that a
glutamate transporter plays a surprising and critical role
at the nonmammalian cone synapse. A further twist appears when we compare the behavior of nonmammalian
and mammalian cones. Thus it is necessary to discuss the
difference.
1. Nonmammals
The Ca2⫹ current in nonmammalian cones appears to
be ill suited for controlling the release of transmitter (see
Fig. 4B). In the absence of light, the membrane voltage is
between ⫺35 and ⫺40 mV. A brief flash produces a hyperpolarization that reaches a maximum of approximately ⫺65 mV and lasts several hundred milliseconds.
The entire presynaptic voltage range contains information
that must be communicated to postsynaptic neurons.
Nonmammalian photoreceptors express a relatively sim1. The voltage required for half-maximum
activation of the voltage-gated Ca2⫹ current
in photoreceptors
TABLE
Species
Rod/Cone
pH
V1/2, mV
Reference Nos.
Salamander
Salamander
Salamander
Salamander
Salamander
Salamander
Turtle
Lizard
Salamander
Salamander
Goldfish
Monkey
Tree shrew
Squirrel
R
R
R
R
C
C
C
C
R
C
C
C
C
C
7.3
7.2–7.3
7.4
7.4
7.4
7.4
7.4
7.4
7.8
7.8
7.8
7.4
7.4
7.4
⫺18 ⫾ 6
⫺22 ⫾ 6
⫺13.1 ⫾ 3.1
⫺17.8
⫺15.3 ⫾ 2.3
⫺19
(⫺20)
(⫺20)
⫺28.2 ⫾ 1.6
⫺26.6 ⫾ 1.1
⫺31
(⫺49)
⫺41.5
(⫺47)
9
25
115
70
115
11
76
87
134
134
150
158
138
34
Values in parentheses have been estimated from data that appear
within figures of the cited references. Otherwise, values have been taken
from the text in each citation. The calcium current in nonmammalian
rods and cones activates at similar voltages. The activation voltage is
shifted approximately ⫺7 mV for a 0.4 increase in pH. The voltage for
half-maximum activation of Ca2⫹ current in mammalian cones is ⬃28
mV more negative than in nonmammalian cones.
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less than several hundred Daltons in molecular mass (33,
see also Ref. 148) and should allow GABA (and other
small molecules, like glutamate and ATP) to leak out of
the cytoplasm and into the synaptic space. Thus hemi-gap
channels might also be sites of transmitter release.
3) The surround signal generated in horizontal cells
is transmitted to cones and hyperpolarizing bipolar cells
at inhibitory synapses and transmitted to depolarizing
bipolar cells at excitatory synapses. It is difficult to see
how ephaptic transmission could produce opposite polarity signals in adjacent cells. In contrast, the polarity of
responses produced by GABA receptors could be determined by differences in intracellular Cl⫺ concentrations
(see Ref. 147).
The existence of hemi-gap junction channels in the
tips of horizontal cell dendrites needs to be confirmed. It
is not clear if a GABA-mediated component of surround
inhibition is eliminated by SKF89976A. The pharmacological specificity of carbenoxolone is uncertain. Nonetheless, the ability of carbenoxolone to inhibit a component
of the surround signal emphasizes that GABA and GABA
transport are not the whole story.
886
E. A. SCHWARTZ
Physiol Rev • VOL
porter) were separately inhibited to determine their individual contributions to controlling the glutamate concentration in the synaptic cleft. As expected, exocytosis
added glutamate to the synaptic cleft. Transporters controlled the rate of removal. The steady-state concentration in the synaptic cleft was determined by a balance
between the ability of exocytosis to add glutamate and the
ability of a transporter to remove glutamate from the
synaptic cleft.
The following experiment demonstrates that the voltage dependence of the transporter was sufficient to control the concentration of glutamate in the synaptic cleft.
The release of glutamate was first blocked with 20 mM
Mg2⫹. As a result, horizontal cells hyperpolarized, and
flashes of light failed to produce synaptic responses (see
also Ref. 38). Next, glutamate was added to the bath and
allowed to diffuse into the synaptic cleft. As expected,
horizontal cells depolarized. But now, the surprising observation was that flashes of light produced relatively
normal responses even though Ca2⫹ entry and exocytosis
remained blocked. Changes in presynaptic voltage (without Ca2⫹ entry) changed the concentration of glutamate
sensed by postsynaptic receptors from 70 to 2 ␮M.
Two factors allow glutamate to act at a relatively low
concentration. First, “on” bipolar cells use metabotropic
receptors, which have a relatively high glutamate sensitivity (see Refs. 121, 130). Second, ionotropic receptors in
horizontal and “off” bipolar cells are desensitized by the
continuous release of glutamate (34, 39). Desensitized
receptors produce small macroscopic currents but, paradoxically, operate with a high glutamate sensitivity (see,
for example, Ref. 50). The transporter modulates the extracellular glutamate concentration within a range that
effectively controls the activation of metabotropic and
desensitized ionotropic receptors.
The voltage-gated Ca2⫹ current is activated in only a
small part of the physiological range, when a cone depolarizes above ⫺40 mV. Within this narrow range, the Ca2⫹
current triggers exocytosis and should accentuate a transient off depolarization that occurs when a light is extinguished (123). The presence of two systems for Ca2⫹
entry, voltage-gated channels and cGMP-gated channels,
may allow two pathways for modulating transmitter release. The relative contribution of the two mechanisms is
uncertain. Indeed, the ratio may not be fixed. Small shifts
in either the voltage dependence of the voltage-gated
Ca2⫹ current or the cGMP concentration may have a
marked effect.
In the standard synapse, a burst of transmitter is
rapidly released and then removed by diffusion and the
steady activity of transporters. Membrane voltage controls Ca2⫹ influx, vesicle fusion, and transmitter release.
Transporters maintain a slow and constant recovery effort. Nonmammalian cones may reverse this situation.
The balance between delivery and removal determines
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ple mix of voltage- and Ca2⫹-dependent currents (9, 87).
The Ca2⫹ current activates when the potential is above
⫺40 mV and has a half-maximum amplitude at approximately ⫺20 mV (Table 1). Hence, there is only a small
overlap between voltages produced during exposure to
light (⫺35 to ⫺65 mV) and voltages that activate the Ca2⫹
current (⫺40 to 0 mV). Two mechanisms have been suggested to resolve the paradox: 1) the voltage dependence
of the Ca2⫹ current is shifted to overlap the functional
range, or 2) Ca2⫹ enters the cytoplasm through another
route.
Various factors have been suggested to modulate the
voltage dependence of the photoreceptor’s Ca2⫹ current:
protons (11), anions (140), divalent cations (106), nitric
oxide (70), dopamine (134), somatostatin (2), and lipids
(149). All of these may produce small shifts (5–10 mV) in
voltage dependence. It is not clear that any agent or
combination can produce the larger shift (20 –30 mV)
needed to align the voltage dependence of the cone’s Ca2⫹
current with the physiological operating range.
Rieke and Schwartz (115) have identified an alternate
path for Ca2⫹ entry. Salamander cones have a cGMPgated conductance in their somata and synaptic endings.
The location of cGMP-gated channels in the synaptic ending has been confirmed by pulling patches from the pedicle of a lizard cone (120). Ca2⫹ enters through cGMPgated channels and triggers exocytosis (measured as an
increase in cell capacitance). Because these channels are
relatively voltage independent within the physiological
range, voltage does not modulate Ca2⫹ entry and exocytosis. Instead, the cGMP-gated conductance provides a
steady influx of Ca2⫹, which supports a continuous exocytosis. The concentration of glutamate in the synaptic
space is controlled by another mechanism.
Salamander cones express a glutamate transporter in
their synaptic endings (105). A series of experiments (44),
described below, indicate that the transporter controls
the concentration of glutamate in the synaptic cleft. These
experiments relied on selectively blocking the activity of
the transporter with dihydrokainate (DHK), blocking
Ca2⫹ entry and exocytosis with extracellular Mg2⫹, and
using light to change the cone’s membrane voltage.
The first experiment determined the relationship between the extracellular glutamate concentration and the
membrane potential of horizontal cells in a retinal slice.
Glutamate was added to the bath after both release (by
exocytosis) and uptake (by the transporter) were
blocked. The membrane potential of horizontal cells depolarized from ⫺70 to ⫺20 mV as the glutamate concentration was changed from 1 to 80 ␮M (EC50 ⫽ 32 ␮M). The
observed correspondence between glutamate concentration and membrane voltage was used in subsequent experiments to translate horizontal cell voltage into the
apparent glutamate concentration in the synaptic cleft.
Release (by exocytosis) and uptake (by the trans-
TRANSPORT-MEDIATED SYNAPSES IN THE RETINA
the concentration of transmitter in the synaptic cleft. A
steady influx of Ca2⫹ through cGMP-gated channels sustains a relatively constant rate of exocytosis (115). The
voltage dependence of transport-mediated uptake appears to be sufficient to control the concentration of
transmitter in the synaptic cleft (44).
2. Mammals
C. Other Synapses?
I have presented the evidence for a transport shuttle
in nonmammalian horizontal cells and transport retrieval
in nonmammalian cones. Does transport-mediated release only occur in slimy animals, viz., amphibians, reptiles, and fish, or is it also important for understanding the
mammalian brain? Our slimy cousins have bigger cells,
which are relatively easy to study. Slow responses in
mammalian dendrites are difficult to study. Transportmediated release is evident in the mammalian brain during pathological conditions and pharmacological intervention. Ischemia releases glutamate by reverse transport
(3). Amphetamines stimulate the reverse transport of dopamine (54). These are interesting phenomena, but the
Physiol Rev • VOL
more important question is the role of transporters in
normal communication. Is there a transport-mediated
synapse in the mammalian brain?
Amacrine cells are retinal interneurons with processes restricted to the inner synaptic layer. Starburst
amacrine cells, named for their distinctive anatomy, are
the only retinal neurons that use acetylcholine as a transmitter. Thus it was puzzling when immunohistochemistry
revealed a high GABA concentration in their cytoplasm
(18, 68, 144). Starburst amacrine cells contain both acetylcholine and GABA. Acetylcholine is stored in synaptic
vesicles, and its release is Ca2⫹ dependent (100). In contrast, GABA is stored predominantly in the cytoplasm, and
its release appears to be Ca2⫹ independent (100). The
mechanism of GABA release is uncertain but may involve
a transporter. An individual neuron might use exocytosis
and transporters to release different transmitters. The
two mechanisms would have different sensitivities to presynaptic voltage and ion concentrations and convey different temporal signals.
And what about the rest of the brain? Are there
transport-mediated synapses outside the retina? The substantia nigra provides an opportunity to briefly discuss
another transmitter, dopamine. Dopaminergic cells in the
pars compacta project axons to the striatum, where they
release dopamine at conventional synapses. Back in the
substantia nigra, the cell bodies and dendrites receive
inputs and also release dopamine (26, 46, 54, 97, 114).
Somatodendritically released dopamine may diffuse tens
of microns (see Ref. 27) before reaching receptors at
nonsynaptic sites (129) and producing an autoinhibitory
feedback (110). The mechanism of release is uncertain.
Although quantal release has been detected at the cell
body (56), electron microscopy reveals few conventional
synapses along dendrites (98). It seems unlikely that the
observed changes in extracellular dopamine could be sustained by a sparse distribution of conventional synapses.
On the other hand, the abundant expression of the dopamine transporter (98) makes reverse transport possible.
Axon terminals and dendrites may use different mechanisms for dopamine release. Release from axon terminals
in the striatum is Ca2⫹ dependent, whereas release from
dendrites in the substantia nigra is largely Ca2⫹ independent (23, 54). Falkenberger et al. (40) have recorded from
cells with patch pipettes and detected extracellular dopamine with amperometry. They blocked the dopamine
transporter with a specific inhibitor and blocked exocytosis by removing extracellular Ca2⫹. An autoinhibitory
current was attributed to reverse transport. Unfortunately, blocking the transporter or removing Ca2⫹ was
slow and required 60 –90 min. During this interval much
can change. It is not clear that the signal seen after
superfusing a slice for 90 min with EGTA also operates in
a normal brain. Thus the role of transport-mediated release remains uncertain. The problem is still to identify
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The Ca2⫹ current in mammalian photoreceptors is
well suited for controlling transmitter release (Fig. 4B,
blue trace). Like other L-type Ca2⫹ currents, it is blocked
by nisoldipine and has little voltage-dependent inactivation. Unlike the L-type currents expressed in cardiac and
somatic muscle, which have a half-maximum activation at
⬃0 mV, the current in mammalian photoreceptors is halfmaximally activated at a relatively hyperpolarized potential, approximately ⫺47 mV (Table 1). The remarkable
leftward shift of Ca2⫹-channel activation in mammalian
photoreceptors may be due to the expression of a unique
channel protein. Genetic and molecular studies have identified a unique L-type channel as the cause of “congenital
stationary night blindness” (CSNB2) (13, 135). The channel protein is expressed in photoreceptors near synaptic
ribbons (91, 92).
Evolution appears to have provided nonmammalian
and mammalian cones with different solutions for the
control of exocytosis and the concentration of transmitter
in the synaptic cleft. Mammalian cones use a unique
voltage-gated Ca2⫹ channel, which activates over the entire physiological voltage range. Synaptic transmission
from cones to bipolar cells is entirely Ca2⫹ dependent
(34). Moreover, there is no indication of a cGMP-gated
conductance in the synaptic ending of squirrel cones (personal observation). In contrast, nonmammalian cones use
a more prosaic voltage-gated channel and supplement
Ca2⫹ entry with a cGMP-gated channel. The voltage dependence of the transporter determines the concentration
of transmitter in the synaptic cleft.
887
888
E. A. SCHWARTZ
the exact contribution of transport-mediated release during physiological conditions.
different tasks. Despite great success, our understanding
of synaptic communication is still rudimentary.
III. QUESTIONS AND CONCLUSIONS
Address for reprint requests and other correspondence: E.
Schwartz, Dept. of NPP, MC0926, Univ. of Chicago, 947 E. 58th
St., Chicago, IL 60637 (E-mail: [email protected]).
Physiol Rev • VOL
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The role that transporters play in transmitter release
has been a topic for 30 years (102, 111, 124). Despite its
long history, relatively few transport synapses have been
identified. Is this because 1) transport synapses are rare,
or 2) transport synapses are difficult to identify and we
are reluctant to acknowledge their role without overwhelming evidence? The past decade has produced the
cloning and electrophysiological characterization of
transport proteins. As a consequence, the mechanism of
transmitter translocation is roughly understood. Now, the
problem is to determine the exact functional role that
transporters play in synaptic communication. Are transporters sometimes important in the moment-to-moment
determination of information flow, or are they always part
of a support staff, which tidies up after a transmitter is
released and corrects small changes in transmitter concentration? Even in the nonmammalian retina, questions
remain concerning the exact role of GABA transporters in
communication by horizontal cells. How do other transmitters and non-GABAergic cells contribute to the surround signal and adaptation?
Why is transport-mediated communication difficult
to study? First, reverse transport must be separated from
the release of transmitter by exocytosis at both synaptic
and nonsynaptic sites. Second, we may be mesmerized by
action potentials, propagation, and rapid communication.
As a consequence, we incorrectly expect transporters to
emulate or participate in similar events. Presently, the
standard synapse can account for communication between most neurons. However, the nervous system may
operate simultaneously on several temporal and spatial
scales. In addition to the rapid transfer of information
from one brain part to another, slower signals may wax
and wane in local regions (see, for example, Ref. 122).
Extracellular recording detects only the timing of action
potentials. Patch pipettes and sharp electrodes usually
record from somata and do not see activity in distant
dendrites, where transport-mediated release may operate.
Our view of brain function may be limited by the same
techniques that have successfully emphasized action potentials, propagation, and rapid communication.
The standard synapse and transport shuttle are models for synaptic function. Standard synapses are optimized for rapid communication. Transport-mediated synapses are expected to operate on a slower time-scale.
Both mechanisms are likely to be embellished and altered
when expressed in real neurons. Real synapses elaborate
features that make them suited for individual tasks. Synapses are not all the same, and individual synapses have
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