TEMPORAL ANGIOTENSIN-INDUCED PHOSPHORYLATION OF ERK1/2 AND OXYTOCIN EXPRESSION IN MALE HYPOTHALAMIC NEURONAL CULTURE.

B. BAEZ; C. PORCARI; VIVAS L.; J. CAMBIASSO ; A. GODINO

Itacuruçá, Río de Janeiro

Congreso; III SMOA- Symposium Miguel Ozorio de Almeida; 2015

Federal Rural University of Río de Janeiro

It is well known that angiotensin II (ANGII) induces thirst and sodium appetite (SA), however when animals are sodium depleted, thirst appears immediately but SA arises with a temporary delay. In vivo experiments have been demonstrated that icv ANGII induce SA behavior due to ERK1/2 phosphorylation at hypothalamic level (Daniels et al., 2009, Felgendreher et al., 2013). The aim of the present study was to analyze in male hypothalamic neuronal cultures the temporal effect of ANGII on induced phosphorylation of ERK1/2 and oxytocin (OT) and angiotensinogen (AOGEN) expression, since these systems have been involved in the inhibition and induction of SA, respectively. Besides we also analyzed whether this ANGII action maybe mediated through angiotensin AT1 receptor (AT1R)We analyzed in primary hypothalamic neuronal cultures (E19) of male rat fetuses after 5DIV the effect of the stimulation of 100nM of ANGII for 15 minutes, 2 and 24 hours. In order to study the participation of AT1R in these effects the cultures were pretreated with the AT1-antagonist, losartan, for 20 minutes before ANGII exposure. After the treatments, the cultures were homogenized and analyzed by western blot for ERK1/2 phosphorylation and the OT and AOGEN expression.We determinate that ANGII rapidly increased ERK1/2 phosphorylation (within 15 min) compared to baseline and those cultures stimulated for a longer time [F(3, 20)=4.58, p=0.13; n=6]. Also, ANGII significantly increases OT expression at 2 hours [F(3, 12)=5.5955, p=0.01233; n=4], relative to the other treatments. We also observed that pre-treatment withlosartan, blocks both effects induced by ANGII (interaction time vs treatment: ERK1/2=F(3, 16)=4.9343, p=0.01297 and OT= F(3, 16)=5.2155, p=0.01057; n=3). The ANG II stimulation did not produce any significant difference on AOGEN expression.In summary, these results provide new information regarding the temporal pattern of ANGII stimulation in hypothalamic neurons on the activity of ERK1/2 and OT expression. These data help to better understand the angiotensin-intracellular signaling pathways, its temporal pattern of activation and the interaction with other systems, such as the oxytocinergic system, during the modulation of thirst and SA.