William J. Brunken

Rod pathways in mammalian retinae

A variety of recent experiments has resolved the way in which signals are transmitted from rod photoreceptors to ganglion cells in the mammalian retina. Rods connect to a single class of rod bipolar cell, which depolarize in response to light. Rod bipolar cells are not connected directly to ganglion cells: they synapse onto rod amacrine cells, which excite ON-centre ganglion cells via gap junctions, and inhibit OFF-centre ganglion cells via inhibitory glycine synapses. Monoamines have particular influences on the rod system, through synapses with rod amacrine and rod bipolar cells, and a function for dopamine and indoleamines within this system can be hypothesized from recent experiments. There is evidence to suggest that dopaminergic amacrine cells bring the surround response into the rod system through synapses with the rod amacrine cell, and that an indoleamine, probably serotonin, increases the signal in the ON pathway through a feedback synapse onto the rod bipolar terminal.

S-laminin expression in adult and developing retinae: A potential cue for photoreceptor morphogenesis

The development of the neural retina follows a stereotyped time course that begins with an undifferentiated neuroepithelium populated by multipotential progenitor cells and ends with a highly differentiated tissue containing diverse cell types. The identities of the factors that guide this differentiation have remained elusive; a likely location for such factors, however, is the extracellular environment. Here, we show that the extracellular matrix component s-laminin is present in the neural retina, that s-laminin expression parallels the differentiation of rod photoreceptors, that photoreceptors interact with s-laminin in vitro, and that antibodies to s-laminin profoundly reduce the appearance of cells that express rhodopsin in vitro. These data suggest that s-laminin plays a role in the differentiation of the neural retina and provide evidence that the composition of the extracellular matrix may be an important determinant of retinal differentiation.

5-HT2a receptors in the rabbit retina: potential presynaptic modulators.

Three 5-HT receptors have been implicated in retinal processing but positive identification of the receptors and the localization of receptor subtypes in the retina have not been achieved. In this study, molecular techniques were used to identify one class of 5-HT receptor--5-HT2a--in the retina, and immunohistochemical techniques were used to localize the receptor in the retinal network. Reverse transcription polymerase chain reaction (RT-PCR) techniques were used to identify a segment of the rabbit 5-HT2a gene; a 422 base fragment was identified, cloned, and sequenced. The fragment shows a high degree (ca. 90%) of nucleotide sequence identity with the 5-HT2a receptor gene from other mammals. 5-HT2a immunoreactivity was seen in both the inner and outer plexiform (synaptic) layers of the retina. Using cell-type-specific markers, the 5-HT2a immunoreactivity was shown to be on the terminals of photoreceptor and rod bipolar cells. This association of 5-HT2a receptors with these two synapses suggests that serotonin may be a modulator of synaptic function in the retina.

Identification of 5-HT3A and 5-HT3B receptor subunits in mammalian retinae: potential pre-synaptic modulators of photoreceptors

Although serotonin (5-HT) is found in the mammalian retina only at low levels, considerable evidence suggests that it plays a role in visual processing. Pharmacological experiments indicate that numerous receptors for 5-HT are present in the mammalian retina. One of these is the ionotropic 5-HT(3) receptor. So far, two subunits for this receptor have been identified in the nervous system, 5-HT(3A) and 5-HT(3B). Co-expression of these subunits in Xenopus oocytes is sufficient to reconstitute native 5-HT(3) receptor properties. Thus, it is believed that a native neuronal 5-HT(3) receptor is multimeric similar to the related acetylcholine receptor family. To determine whether this receptor is expressed in the mammalian retina, we first performed reverse transcription polymerase chain reaction and first demonstrated the presence of transcripts for both the 5-HT(3A) and 5-HT(3B) receptor subunits. Then using a well-characterized polyclonal antiserum against the 5-HT(3A) receptor subunit, we demonstrated 5-HT(3A) receptor immunoreactivity (IR) in the rabbit, rat, and human retina. This IR was localized specifically to the rod photoreceptor terminals in all three species, suggesting that this receptor may modulate the rod signaling pathway by controlling the output at the rod terminals.

Roles of the extracellular matrix in retinal development and maintenance.

The extracellular matrix (ECM) is a highly organized meshwork of secreted macromolecules that consists of proteins, proteoglycans, and polysaccharides. Components of the ECM provide cells with different signals that are involved in many aspects of the development of multicellular organisms, such as: (1) when to start, continue, or stop dividing; (2) when to differentiate; (3) where to migrate; (4) how to polarize; (5) where to form a synapse; and (6) whether to die or survive. Thus, ECM molecules may be playing many different roles in the development of a complex tissue such as the retina.

The Function of Monoamines in the Rabbit Retina

Over the last several years we have described the action of dopamine and indoleamine drugs on the activity of ganglion cells in the rabbit retina in the mesopic state (Jensen & Daw, 1984, 1986; Brunken & Daw, 1987, 1988a, 1988b). The prime action of dopamine is that it reduces the sensitivity of both ON and OFF responses, while D1 antagonists reduce or abolish the surround responses in both ON and OFF center cells. The prime action of serotonin is that it increases spontaneous activity and responses in ON cells, while 5HT2 antagonists have an opposite effect. For both transmitters there is a second receptor with actions opposite to the firsts: D2 receptors with actions opposite to D1, and 5HT1A receptors with actions opposite to 5HT2• There are other effects which will be described below, but these are the prime ones.

A differential effect of APB on ON- and OFF-center ganglion cells in the dark adapted rabbit retina.

The glutamate analog, 2-amino-4-phosphonobutyric acid (APB) is a proven tool in exploring the retinal circuit; it has been shown to interfere specifically with the transmission from photoreceptor to depolarizing bipolar cell. Consequently, in photopic retinae, the application of APB disrupts the ON-channel leaving the OFF-channel undisturbed; on the other hand, in the scotopic state, APB application blocks all ganglion cell responses. In this paper, we will show that the ON- and OFF-channels have a differential sensitivity to application of APB. That is to say, APB blocks center responses in ON-ganglion cells at mean concentration of 22 +/- 5.1 microM (mean +/- standard error of the mean; n = 15) and in OFF-ganglion cells at mean concentration of 91 +/- 15.5 microM (n = 16). Since considerable data rule out direct effects of APB on ganglion cells, we hypothesize that this effect is due to a difference in the synaptic gain of ON and OFF pathways in the inner retina.