Effect of Dp71 deficiency on the oxytocin hypothalamic axis in osmoregulation function in mice.

Dp71 is the major form of dystrophins (Dp) in the supraoptic nucleus (SON) and in the neural lobe of hypophysis (NL/HP). Dp71-null mice exhibit a hypo-osmolar status attributed to an altered osmosensitivity of the SON and to a perturbed vasopressinergic axis. Because oxytocin (OT) is implicated in osmoregulation via natriuresis, this study explored the oxytocinergic axis in Dp71-null mice after salt-loading (SL). Under normosmolar conditions, OT-mRNA expression was higher in the Dp71-null SON compared to wild-type (wt) and the OT peptide level has not changed. Dp-immunostaining was localized in astrocytes end-feet surrounding vessels in wt SON. This distribution changed in Dp71-null SON, Dp being detected in OT-soma of MCNs. nNOS and NADPH-diaphorase levels increased in the OT area of the Dp71-null SON compared to wt. In the NL/HP, OT level reduced in Dp71-null mice and Dp localization changed from pituicytes end-feet in wt SON to OT terminals in Dp71-null SON. Salt-Loading resulted in an increase of OT-mRNA and peptide levels in wt SON but had no effect in Dp71-null SON. In the NL/HP, OT content was reduced after SL. For Dp71-null mice, OT level, already low in control, was not modified by SL. Dp level was not affected by SL in the SON nor in the NL/HP. Our data confirmed the importance of Dp71 for the SON functionality in osmoregulation. The localization of Dp71 at the glial-vascular interface could be associated with SON osmosensitivity, leading to an adequate OT synthesis in the SON and release from the NL/HP upon plasmatic hyperosmolality.

Mammalian retinal horizontal cells are unconventional GABAergic neurons.

Lateral interactions at the first retinal synapse have been initially proposed to involve GABA by transporter-mediated release from horizontal cells, onto GABA(A) receptors expressed on cone photoreceptor terminals and/or bipolar cell dendrites. However, in the mammalian retina, horizontal cells do not seem to contain GABA systematically or to express membrane GABA transporters. We here report that mouse retinal horizontal cells express GAD65 and/or GAD67 mRNA, and were weakly but consistently immunostained for GAD65/67. While GABA was readily detected after intracardiac perfusion, it was lost during classical preparation for histology or electrophysiology. It could not be restored by incubation in a GABA-containing medium, confirming the absence of membrane GABA transporters in these cells. However, GABA was synthesized de novo from glutamate or glutamine, upon addition of pyridoxal 5'-phosphate, a cofactor of GAD65/67. Mouse horizontal cells are thus atypical GABAergic neurons, with no functional GABA uptake, but a glutamate and/or glutamine transport system allowing GABA synthesis, probably depending physiologically from glutamate released by photoreceptors. Our results suggest that the role of GABA in lateral inhibition may have been underestimated, at least in mammals, and that tissue pre-incubation with glutamine and pyridoxal 5'-phosphate should yield a more precise estimate of outer retinal processing.

A deficit of brain dystrophin 71 impairs hypothalamic osmostat.

Patients with Duchenne muscular dystrophy (DMD) and mdx mice, devoid of dystrophin proteins, show altered ionic homeostasis. To clarify dystrophin's involvement in the central control of osmotic stimuli, we investigated the effect of the disruption of Dp71, the major form of dystrophin in the brain, on the hypothalamoneurohypophysis system (HNHS) osmoregulatory response. Dp71 and Dp140 are the principal DMD gene products in the supraoptic nucleus (SON) and neurohypophysis (NH). They are present in astrocyte and pituicyte end-feet, suggesting involvement in both intrinsic osmosensitivity of the SON and vasopressin (AVP) release from the NH. In Dp71-null mice, the cellular distribution of Dp140 was modified, this protein being detected on the membrane of magnocellular soma. The plasma osmolality of Dp71-null mice was lower than that of wild-type mice under normal conditions, and this difference was maintained after salt loading, indicating a change in the set point for osmoregulation in the absence of Dp71. The increase in AVP levels detected in the SON and NH of the wild-type was not observed in Dp71-null mice following salt loading, and the increase in AVP mRNA levels in the SON was smaller in Dp71-null than in wild-type mice. This suggests that Dp71 may be involved in the functional activity of the HNHS. Its astrocyte end-feet localization emphasizes the importance of neuronal-vascular-glial interactions for the central detection of osmolality. In the SON, Dp71 may be involved in osmosensitivity and definition of the "osmostat," whereas, in the neurohypophysis, it may be involved in fine-tuning AVP release.