Retinoic acid receptor (RAR)-alpha is not critically required for mediating retinoic acid effects in the developing mouse retina.

PURPOSE: To determine the functional contribution of retinoic acid receptor (RAR)-alpha in the developing murine neural retina, through a phenotypic analysis of the corresponding null mutants. METHODS: RARalpha mutant (Rara(-/-)) mice were compared with wild-type littermates at several stages of pre- and postnatal development. An RA-response element (RARE)-containing reporter transgene was used to assess the contribution of RARalpha to retinoid signaling in the retina. In situ hybridization was performed on serial eye sections to investigate the expression of main developmental regulators. Immunofluorescence was used to detect differentiated cell types in the adult retina. Mutants were also subjected to clinical observation and visual function evaluation with the optomotor test and electroretinography. RESULTS: Both isoform transcripts of RARalpha were expressed throughout the neural retina at various stages of pre- and postnatal development. In the Rara(-/-) mice the RARE-reporter transgene consistently failed to activate in the developing neural retina. However, they did not exhibit any alteration of the expression patterns of molecular determinants and had a normal organization of retinal cell types at postnatal stages. Their performance in visual tests was indistinguishable from that of control littermates. CONCLUSIONS: Although RARalpha mediates RARE reporter transgene activity in the neural retina, its function is not necessary for the retina to develop and function normally. These data suggest that retinoic acid regulates neural retinal development through other, possibly RAR-independent, pathways.

Functional evidence for a role of GABA receptors in modulating nerve activities of circular smooth muscle from rat colon in vitro.

In the enteric nervous system, activation of neuronal GABA(A)- and GABA(B)-receptors has been shown to modulate neuronal activity. The consequences of this modulation depend on the location in the gastrointestinal tract or the animal species studied. These data illustrate the complexity of GABA-induced effects. Furthermore, the GABA(C)-receptor has been identified in a neuroendocrine cell line suggesting a modulating role of this third type of GABA receptor in intestinal functions. Therefore, the modulating role of GABA-receptor agonists was determined in circular preparations of rat distal colon during electrical nerve stimulation (NS) in vitro. Mechanical response to NS was characterized by a relaxation followed at the end of the stimulation by an off-contraction. In normal Krebs solution (basal conditions), muscimol and baclofen, respectively GABA(A)- and GABA(B)-agonists, induced a significant increase of the electrically induced off-contraction. The GABA(C) agonist, CACA, showed no significant effect on the response to NS. Excitatory effects of muscimol on the off-contraction were abolished in the presence of atropine. Furthermore, in the presence of atropine, muscimol increased the amplitude of the electrically induced relaxation; similarly the baclofen-induced increase of off-contraction amplitude was significantly lower than that observed in control conditions. Baclofen and muscimol effects on the off-contraction were abolished in the presence of hexamethonium or guanethidine. Furthermore, muscimol and baclofen did not induce any significant change on the response to NS in the presence of L-NAME and apamin together. Thus, it seems that in rat distal colon, GABA regulates significantly both excitatory (through GABA(A)- and GABA(B)-receptors) and inhibitory (through GABA(A)-receptors) neuronal activities. We also gave evidence for a possible interplay between GABAergic intrinsic neurons and adrenergic nerve terminals. Finally, it is shown for the first time the presence of the GABA vesicular transporter (VIAAT) around myenteric ganglia of rat colon.

Cellular localization of the vesicular inhibitory amino acid transporter in the mouse and human retina.

Horizontal cells are classically thought to mediate lateral inhibition by gamma-aminobutyric acid (GABA)-transporter mediated release. In the mammalian retina, however, GABA uptake and cloned GABA transporter were not detected in horizontal cells. Furthermore, the vesicular inhibitory amino acid transporter (VIAAT or VGAT) that loads GABA and glycine into synaptic vesicles was reported recently to be expressed in horizontal cells. To further assess synaptic transmission in mammalian horizontal cells, we examined the subcellular distribution of VIAAT in mouse and human retina by confocal microscopy with specific cell markers. VIAAT was observed in the mouse outer plexiform layer as punctate structures that localized in calbindin-positive horizontal cells. These structures were in close apposition with synaptophysin-, PSD-95-, dystrophin-, and bassoon-immunopositive photoreceptor terminals, suggesting that VIAAT is localized in horizontal cell tips at photoreceptor terminals. VIAAT-positive puncta were also in apposition to lectin-labeled cone terminals or dendrites of PKCalpha-immunopositive rod bipolar cells, indicating that VIAAT is expressed in horizontal cell tips at both rod and cone terminals. By contrast, only a very few puncta were observed in the human outer plexiform layer, whereas the inner plexiform layer remained labeled as in the mouse retina. When using adult human retinal cells in culture, horizontal cells identified by parvalbumin immunostaining were found to contain VIAAT, either at their terminals or throughout the entire cell similarly as in syntaxin-immunopositive cells. These differences between human retinal tissue and cultured cells were attributed to VIAAT degradation in postmortem retinal tissue. VIAAT localization in mouse and human horizontal cells further support the role of inhibitory transmitters in lateral inhibition at the photoreceptor terminals.