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Biomedical subjects

D M Lam

Publications and source records attributed to D M Lam.

At least 73 records · Page 4Linked to original sources
PubMed1984

The identification and some functions of GABAergic neurons in the proximal retina of the catfish.

Putative GABAergic neurons in the inner retina of the Texas channel catfish, Ictalurus punctatus, were studied using autoradiographic, biochemical and electrophysiological techniques. A pyriform amacrine cell, ramifying predominantly in sublamina b of the inner plexiform layer, was found to accumulate exogenous GABA in the dark. GABA could be released from these cells with high K+ Ringers solution and the release was blocked by Co2+. These data suggest that a class of catfish amacrine cells are GABAergic. To substantiate this, extracellular recordings were made from ganglion cells during the superfusion of the GABA blocking agent bicuculline methochloride. This agent modified the responses of ganglion cells in a manner consistent with the notion that GABAergic amacrine cells are a major source of input to on-center ganglion cells. There also appeared to be an indirect GABAergic influence on a subclass of off-center ganglion cell.

Action Potentials↗
PubMed1984

Visual cycle in the mammalian eye. Retinoid-binding proteins and the distribution of 11-cis retinoids.

This work was designed to provide an insight into the mammalian visual cycle by investigating the possible function of retinoid-binding proteins in this system, and the distribution and type of 11-cis retinoids present in the interphotoreceptor matrix and the cytosols of the retinal pigment epithelium and retina. The total retinol and retinal in the soluble fractions from these three compartments was 8% (3.31 nmol/eye) of the retinyl palmitate and stearate stored in the pigment epithelium membrane fractions (39 nmol/eye). Only small amounts of retinoids were detected in the rod outer segment cytosol. The insoluble fractions also contained retinol, nearly all of which was found in the retina. The retinoids in the soluble fractions appeared to be bound to cellular retinol-binding protein (CRBP), cellular retinal-binding protein (CRA1BP) and interstitial retinol-binding protein (IRBP, a high-Mr glycoprotein). Using immunospecific precipitation, immunoblot and immunocytochemical techniques it was demonstrated that IRBP was localized in the interphotoreceptor matrix and was synthesized and secreted by the retina, a process that did not require the protein to be glycosylated. The amount of retinol bound to IRBP increased if the eyes were exposed to light, when it was estimated that the protein carried up to 30% of its full capacity for all-trans retinol. In addition to all-trans retinol, IRBP carried smaller amounts of 11-cis retinol. The proportion of 11-cis retinol was frequently higher in eyes that had been protected from illumination, suggesting that IRBP plays a role in rhodopsin regeneration during dark-adaptation. Additionally, endogenous 11-cis retinoids in the retina and RPE cytosols were bound to an Mr 33,000 protein tentatively identified as CRA1BP. The 11-cis retinoid in the retina cytosol was mainly in the form of retinol, while in the RPE cytosol it was mainly in the form of retinal. Substantial amounts of 11-cis retinol were also found in the insoluble (membrane) fraction from the retina. It is suggested that in the mammalian retina 11-cis retinol is generated from all-trans retinol (possibly in the Muller cells). Lack of an 11-cis retinol oxidoreductase in the retina prevents it from being utilized for rhodopsin regeneration until it has been transported to the pigment epithelium, where it is converted to 11-cis retinal and returned to the rod outer segments. It is also suggested that IRBP may be implicated in the transport of retinoids between the rod outer segments, the Muller cells and the pigment epithelium.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1984

An extracellular retinol-binding glycoprotein in the eyes of mutant rats with retinal dystrophy: development, localization, and biosynthesis.

Interstitial retinol-binding protein (IRBP) is a soluble glycoprotein in the interphotoreceptor matrix of bovine, human, monkey, and rat eyes. It may transport retinol between the retinal pigment epithelium and the neural retina. In light-reared Royal College of Surgeons (RCS) and RCS retinal dystrophy gene (rdy)+ rats, the amount of IRBP in the interphotoreceptor matrix increased in corresponding proportion to the amount of total rhodopsin through postnatal day 22 (P22). In the RCS-rdy+ rats, the amount increased slightly after P23. However, in the RCS rats there was a rapid fall in the quantity of IRBP as the photoreceptors degenerated between P23 and P29. No IRBP was detected by immunocytochemistry in rats at P28. The amount of rhodopsin fell more slowly. Although retinas from young RCS and RCS-rdy+ rats were able to synthesize and secrete IRBP, this ability was lost in retinas from older RCS rats (P51, P88) but not their congenic controls. The photoreceptor cells have degenerated at these ages in the RCS animals, and may therefore be the retinal cells responsible for IRBP synthesis. The putative function of IRBP in the extracellular transport of retinoids during the visual cycle is consistent with a defect in retinol transport in the RCS rat reported by others.

Aging↗
PubMed1984

Characterization, localization, and biosynthesis of an interstitial retinol-binding glycoprotein in the human eye.

Human eyes contain an Mr 135K retinol-binding protein that is analogous to interstitial retinol-binding protein ( IRBP ) in the subretinal space of bovine eyes. It is a glycoprotein, because it binds 125I-concanavalin A, 125I-wheat germ agglutinin and 125I-Lens culinaris hemagglutinin. It does not bind Ricinus communis agglutinin I. After desialation, it binds Ricinus communis agglutinin I, but loses its capacity to bind wheat germ agglutinin. These observations, coupled with the known specificities of these lectins, suggest that at least one of the oligosaccharide chains is a sialated , biantennary complex type containing fucose. Both by direct analysis of dissected ocular tissues and by immunocytochemistry it was shown that human interstitial retinol binding protein is an extracellular protein that is confined predominantly to the subretinal space. Monkey retinas incubated in vitro in medium containing [3H]leucine were shown to synthesize and secrete this protein into the medium, a conclusion that was confirmed by immunoprecipitation with an immunoglobulin fraction prepared from rabbit antibovine IRBP serum. Virtually no other labeled proteins were detectable in the medium. It is concluded that interstitial retinol-binding protein meets many of the requirements for a putative transport protein implicated in the transfer of retinol between the pigment epithelium and retina during the visual cycle, and that the neural retina may play an important role in regulating its amount in the subretinal space.

Amino Acids↗
PubMed1984

The release of gamma-aminobutyric acid from horizontal cells of the goldfish (Carassius auratus) retina.

Isolated horizontal cells from goldfish retinas were prepared by enzymatic dissociation using papain and separated from other cells by velocity sedimentation. In the intact retina, H1 horizontal cells possess a high-affinity mechanism for accumulating gamma-aminobutyric acid (GABA). This property is retained in isolated cells, which also release the accumulated GABA in response to depolarization by elevated external K+. L-Glutamic acid and its analogues are highly effective at micromolar concentrations in eliciting the release of preloaded GABA from isolated cells. At saturating concentrations, L-aspartic acid stimulates about one-third as much release as L-glutamic acid. In contrast, the D-isomers of glutamate and aspartate are ineffective. In the intact retina, micromolar concentrations of L-glutamic acid analogues are also capable of eliciting GABA release from H1 horizontal cells. Release of the accumulated GABA from isolated H1 cells is largely independent of external Ca2+ concentrations. In the intact retina, H1 horizontal cells also possess a K+-stimulated GABA release mechanism that is independent of the Ca2+ concentrations in the medium. In addition, there appears to be a small but significant amount of [3H]GABA release that may be Ca2+ dependent. Under our conditions, [3H]GABA release from isolated cells is unaffected by external Na+ concentrations between 20 and 120 mM. However, concentrations of 10 mM or less significantly diminishes this release, with 70% curtailed in Na+-free solutions. Our results, together with morphological observations by a number of other investigators, suggest that there may be two distinct mechanisms for GABA release from goldfish H1 horizontal cells: one being a conventional vesicular mechanism which is Ca2+ dependent, while the other is Na+ driven and Ca2+ independent. H1 horizontal cells in the intact goldfish retina release the accumulated GABA in response to brief incubations in darkness, which is known to be the natural stimulus that depolarizes these neurones.

Animals↗
PubMed1983

The ultrastructure of rat rod synaptic terminals: effects of dark-adaptation.

The ultrastructure of synaptic terminals of rat rod photoreceptors was studied in light-adapted and dark-adapted states. In the light-adapted state, horizontal cell processes embedded in the photoreceptor terminal show smooth surface contours and are ovoid in cross section. In the dark-adapted state, fingerlike protrusions of photoreceptor cytoplasm extend into the horizontal cell processes on either side of the synaptic ribbon. An electron-dense structure lies under the horizontal cell membrane that surrounds these fingerlike protrusions. This submembrane specialization is generally associated with areas of concavity of the horizontal cell membrane; it occurs in goldfish rods as well as rat rods. The protrusions and the horizontal cell specialization may be involved in membrane dynamics during synaptic activity.

Animals↗
PubMed1983

Biochemical and biophysical studies of isolated horizontal cells from the teleost retina.

Neurons in the vertebrate retina are interconnected by a complex network of chemical and electrical synapses which make it difficult to study the intrinsic biochemical and biophysical properties of individual cells. During the past decade, enzymatic methods have been developed for dissociating adult retinas into viable and readily identifiable single cells. Using this preparation, we and other investigators have studied the physiological and biochemical properties of several classes of isolated retinal cells. In particular, we have shown that a type of retinal interneuron, the GABA-ergic horizontal cell of teleost retinas, may be an excellent model system for examining the intrinsic membrane properties as well as the cellular mechanisms regulating GABA uptake, synthesis and release from an identified CNS neuron.

Animals↗
PubMed1983

Retinal regeneration in the adult newt, Notophthalmus viridescens: appearance of neurotransmitter synthesis and the electroretinogram.

The removal or destruction of retina in the adult newt, Notophthalmus viridescens, leads to the formation of a new functional retina by metaplasia of the pigment epithelium. In order to understand the physiological and biochemical changes that occur during regeneration, we have examined the time-course of appearance and maturation of the electroretinogram (ERG) and correlated it with the synthesis of the neurotransmitters, acetylcholine (ACh) and gamma-aminobutyric acid (GABA). In addition, we have also determined the pattern of DNA, RNA, and protein synthesis in the regenerating retina. Our experimental results show that: (1) the major peaks of DNA, RNA, and protein synthesis occur at different times during regeneration and that the synthesis of RNA precedes that of DNA which is followed by the protein; (2) synthesis and accumulation of ACh and GABA is lost by the fifth day following ocular devascularization and it reappears by day 23-24. ACh and GABA syntheses continue to increase until they attain plateau around day 35; (3) the appearance of choline acetyltransferase and glutamate decarboxylase activities parallel ACh and GABA synthesis; and (4) transmitter synthesis precedes development of the ERG. The first light evoked response is seen around day 29 by which time the transmitter synthesis is well advanced.

Animals↗
PubMed1983

L-glutamic acid: a neurotransmitter candidate for cone photoreceptors in human and rat retinas.

We have combined immunocytochemical localization of L-aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1; glutamic-oxaloacetic transaminase) with autoradiographic localization of high-affinity uptake sites for L-glutamate or L-aspartate to identify the neurotransmitters of mammalian photoreceptors. In both human and rat retinas, high aspartate aminotransferase immunoreactivity is found in cones but not in rods; certain putative bipolar and amacrine cells are also heavily stained. In the human retina, and perhaps also in the rat retina, cones possess a high-affinity uptake mechanism for L-glutamate but not L-aspartate, whereas rods and Müller (glial) cells take up both L-glutamate and L-aspartate. Taken together, our results indicate that (i) L-glutamate is much more likely than L-aspartate to be the transmitter for human cones, and possibly for cones of other mammalian species as well, and (ii) major differences exist between mammalian cones and rods in the transport and metabolism or utilization of L-aspartate and L-glutamate.

Animals↗
PubMed1982

Dopaminergic neurons in the human retina.

The utilization of dopamine in the adult human retina was examined by using high-affinity uptake, localization, synthesis, and release as neurotransmitter-specific physiological probes. Autoradiographic and histochemical studies have shown that dopamine-accumulating and dopamine-containing cells of the human retina belong to a population of neurons whose somata are located in the proximal regional of the inner nuclear layer. Some of these are amacrine cells which are pre- and postsynaptic to other amacrine cells exclusively in the inner plexiform layer. However, evidence is presented which indicates the existence of interplexiform dopaminergic neurons which send processes to both plexiform layers of the retina. These neurons contain a high concentration of dopamine, take up 3H-dopamine by a hig-affinity mechanism, and release endogenous or accumulated dopamine by a Ca2+-dependent mechanism upon depolarization with high extracellular K+. An endogeneous level of about 20 pmoles dopamine per mg protein was measured in freshly isolated retina using high-pressure liquid chromatography with electrochemical detection. These results demonstrate that mechanisms for dopaminergic neurotransmission are present in the human retina.

Adolescent↗
PubMed1982

Prenatal development of gabaergic, glycinergic, and dopaminergic neurons in the rabbit retina.

The main findings from our studies on the emergence and maturation of neurotransmitter systems in the rabbit retina are that, during normal development: (1) the commitments for certain neurons to become GABAergic, glycinergic, or dopaminergic are made prenatally; the maturation of these neurons, however, occurs postnatally; (2) specific accumulations of exogenously supplied transmitters by presumed retinal neurons were first observed autoradiographically around embryonic day 22 (E22) for gamma-aminobutyric acid (GABA), E25 for glycine, and E27 for dopamine, suggesting that putative GABAergic, glycinergic, and dopaminergic neurons are determined phenotypically at least by these days; (3) based upon the three transmitter-specific properties--uptake, synthesis, and release--the uptake property is generally the first to emerge, while the appearance of the other properties follows a precise and distinct temporal pattern for each of the transmitters studied; (4) for GABAergic and glycinergic systems, the emergence of K+-stimulated, Ca2+-dependent release mechanisms occurs many days after the first appearance of the uptake properties; in contrast, the mechanisms for dopamine uptake and release emerge at about the same time; and (5) by our criteria, putative GABAergic and glycinergic amacrine cells are mature between postnatal day 10 (P10) and P12, around the time when the eyes first open and direction-selective ganglion cells can first be recorded; however, the dopaminergic neurons probably are not mature until about P24.

Animals↗
PubMed1981

Lectin receptors on cells isolated from the turtle retina.

The presence of specific oligosaccharides on the surface of retinal cells was examined by incubating FITC-labeled lectins with cells dissociated from papain-treated turtle retinas. The pattern and intensity of binding was found to vary among the cells examined. With Con A, there was strong surface staining of both rods and cones, with an intense ring of fluorescence above the nucleus. The bipolar and ganglion cells also showed strong surface labeling. In Müller (glial) cells there was intense fluorescence in the apical, microvillous region. In contrast, the horizontal cells and their axons showed weak staining. When RCA-60, RCA-120, and WGA were incubated with photoreceptors, bipolar cells, or horizontal cells, little fluorescence was visible. However, all three lectins bound strongly to the Müller cells. In contrast, the Lotus lectin did not bind to any of the cells examined. In all the cases, lectin binding was inhibited by the appropriate haptene sugar. Further, prior treatment cells with neuraminidase did not alter lectin binding to any cell type. These results suggest differences in the distribution of lectin receptors among specific cell types, and particularly between neurons and glial cells in the vertebrate retina.

Animals↗
PubMed1981

An opiate system in the goldfish retina.

Recently, in addition to conventional neurotransmitters such as acetylcholine, dopamine, glycine and gamma-aminobutyric acid (GABA), putative neuroactive peptide transmitters have been localized to specific retinal amacrine cells. In particular, opiate receptors 2,3, assayable enkephalin immunoreactivity and enkephalin-immunoreactive neurones 1,5 have been described in avian and mammalian retinae. However, little physiological evidence has been obtained for the involvement of neuropeptides in retinal function. Here we report that exogenous opiates affect both the release of GABA from GABAergic amacrine cells and the firing patterns of ganglion cells in the goldfish retina. Our results show that the output of the retina is modulated by an opiate system whose neural organization and pharmacological properties resemble those described elsewhere in the vertebrate central nervous system.

Action Potentials↗
PubMed1981

The emergence, localization, and maturation of neurotransmitter systems during development of the retina in Xenopus laevis: II. Glycine.

The high-affinity uptake and release of glycine was studied in retinas of Xenopus laevis. In the toad and tadpole retina, 3H-glycine was accumulated by a population of cells located predominantly in the inner nuclear layer. When retinas preloaded with 3H-glycine were subjected to high K+-concentrations, these retinas released large amounts of 3H-glycine by a Ca++-dependent mechanism. The appearance and maturation of these putative glycinergic properties was followed during retinal development. Our results indicate that the high-affinity uptake of glycine first appears around stage 33/34 whereas K+-stimulated glycine release cannot be detected until stage 42.

Animals↗
PubMed1981

The emergence, localization, and maturation of neurotransmitter systems during development of the retina in Xenopus laevis. III. Dopamine.

The uptake, synthesis, and release of dopamine was studied in retinas of Xenopus laevis. In the tadpole and adult retina, 3H-dopamine is accumulated by cells located in the inner nuclear layer. Retinas preloaded with 3H-dopamine release this compound in response to high K+ concentrations in the medium. This release is probably Ca++-dependent as it is inhibited by Co++ in the medium. Adult retinas are also capable of synthesizing 3H-dopamine from 3H-tyrosine. The appearance and maturation of these dopaminergic properties were followed during retinal development. Our data indicate that synthesis of dopamine can first be detected as early as stage 35/36 whereas uptake of dopamine first occurs at stage 43. K+-stimulated release of preloaded 3H-dopamine from putative dopaminergic neurons is, however, not evident until stage 46. These results show that similar to the development of GABA-ergic and glycinergic properties, the uptake, synthesis, and release mechanisms for dopamine emerge at different stages during retinal differentiation in Xenopus Laevis.

Acetylcholine↗