High-affinity neurotensin receptor is involved in phosphoinositide turnover increase by inhibition of sodium pump in neonatal rat brain.

Phosphoinositide (PI) metabolism is enhanced in neonatal brain by activation of neurotransmitter receptors and by inhibition of the sodium pump with ouabain or endogenous inhibitor termed endobain E. Peptide neurotensin inhibits synaptosomal membrane Na(+), K(+)-ATPase activity, an effect blocked by SR 48692, a selective antagonist for high-affinity neurotensin receptor (NTS1). The purpose of this study was to evaluate potential participation of NTS1 receptor on PI hydrolysis enhancement by sodium pump inhibition. Cerebral cortex miniprisms from neonatal Wistar rats were preloaded with [(3)H]myoinositol in buffer during 60 min and further preincubated for 0 min or 30 min in the absence or presence of SR 48692. Then, ouabain or endobain E were added and incubation proceeded during 20 or 60 min. Reaction was stopped with chloroform/methanol and [(3)H]inositol-phosphates (IPs) accumulation was quantified in the water phase. After 60-min incubation with ouabain, IPs accumulation values reached roughly 500% or 860% in comparison with basal values (100%), if the preincubation was omitted or lasted 30 min, respectively. Values were reduced 50% in the presence of SR 48692. In 20-min incubation experiments, IPs accumulation by ouabain versus basal was 300% or 410% if preincubation was 0 min or 30 min, respectively, an effect blocked 23% or 32% with SR 48692. PI hydrolysis enhancement by endobain E was similarly blocked by SR 48692, being this effect higher when sample incubation with the endogenous inhibitor lasted 60 min versus 20 min. Present results indicate that PI hydrolysis increase by sodium pump inhibition with ouabain or endobain E is partially diminished by SR 48692. It is therefore suggested that NTS1 receptor may be involved in cell signaling system mediated by PI turnover.

NMDA receptors contribute to the survival promoting effect of high potassium in cultured cerebellar granule cells.

The present report further analyzes the survival promoting effect of high potassium, a condition that mimics neural activity in cultured cerebellar granule cells, an excellent model to study trophic mechanisms induced by depolarization and trophic factors. We found that the survival promoting effect measured at 7 days in vitro (DIV 7) of depolarizing potassium concentrations (25 mM KCl), added at DIV2, is partially prevented by adding at DIV 2 the non-competitive NMDA blocker MK801 (10 microM). The concentration of MK801 used blocks completely the survival promoting effect of a supramaximal effective concentration of NMDA (100 microM). The addition at DIV 2 of anti-brain derived neurotrophic factor (anti-BDNF) antibody, failed to modify the effect of high potassium. The present report provides evidences that in cultured cerebellar granule cells, high potassium-induced survival promoting effect is due in part by the activation of NMDA receptors. The effect does not require the presence of BDNF.