Heterogeneous co-localization of AT 1A receptor and Fos protein in forebrain neuronal populations responding to acute hydromineral deficit

The present study investigates co-localization of AT 1A receptor subtype and Fos protein in neuronal populations of the lamina terminalis (LT) that have been recruited during acute Na + and water depletion mediated by furosemide injections. For that purpose, we combined high cellular resolution of in situ hybridization technique to reveal neurons expressing AT 1A receptor gene (AT 1A mRNA) with the specificity of Fos protein immunoreactivity as a marker of neuronal activation (Fos-ir). As expected, furosemide treatment dramatically increased the density of Fos-immunoreactive neuronal population in all the regions of the LT compared to control (saline-injected animals). Distribution analysis of Fos-ir neurons and AT 1A receptor-expressing neurons performed consecutively to furosemide-induced Na + and water depletion indicated that double-labeled neurons (AT 1A mRNA+Fos-ir) represented the majority (67%) of the neuronal population that expressed AT 1A receptor in the rim of the vascular organ of the lamina terminalis (OVLT). Double-labeled neurons amounted about 60% of the neurons that expressed AT 1A receptor in the core of the subfornical organ (SFO) and 34% in the periphery of the SFO. In the median preoptic nucleus (MnPO), the density of the double-labeled neuronal population observed in the furosemide-treated animals remained weak compared to the control group of animals. Double-labeled neuronal population estimated in the MnPO of the furosemide-treated group of animals represented 17% of the neurons that express AT 1A receptor gene. Our results report a heterogeneous distribution of the neuronal populations that co-localize AT 1A receptor and Fos protein in the lamina terminalis after an acute Na + and water depletion. This study gives anatomical support to a direct action of endogenous AngII on c-fos transcription via binding on AT 1A receptor in specific areas of the circumventricular organs (rim of the OVLT and core of the SFO). In the MnPO, our data indicate that intracellular signaling pathways unlikely couple AT 1A receptor with c-fos transcription. The expression of Fos protein in this nucleus might be therefore secondary to the recruitment of excitatory inputs different from AngII. This observation underlines the complexity of molecules and neurocircuits in the preoptic region that are involved in the control of acute Na + and water deficit.