Phospholipase A₂ and Somatostatin Release are Activated in Response to N‐Methyl‐D‐Aspartate Receptor Stimulation in Hypothalamic Neurons in Primary Culture

Abstract

We have recently shown that glutamate primarily induces somatostatin release in hypothalamic neurons through N-methyl-Daspartate (NMDA)-type receptor sites. Here we report that glutamate and NMDA also stimulate the release of [3H]arachidonic acid in a dose-dependent manner. The NMDA-induced effects (arachidonic acid release and somatostatin secretion) were both inhibited by MK-801, an NMDA receptor-type antagonist, or mepacrine, a phospholipase A, inhibitor. In addition, mepacrine was able to inhibit A23187-stimulated arachidonic acid release and somatostatin secretion. p-Bromophenacylbrornide, another phospholipase A, inhibitor, also blocked NMDA-induced secretion of somatostatin. However, responses to NMDA were unaffected by H7 (inhibitor of protein kinase C), nordihydroguaiaretic acid or indomethacin (inhibitors of lipoxygenase and cyclooxygenase). Melittin, a phospholipase A, activator, was found to stimulate both responses, but omission of extracellular CaZ from the incubation media strongly reduced melittin-induced somatostatin release. Six-h pertussis toxin pretreatment did not significantly reduce the action of NMDA on either of the two parameters studied. High-performance liquid chromatography analysis of [3H]metabolites released in the medium after NMDA stimulation revealed that [3H]arachidonic acid was the only detectable metabolite. External addition of arachidonic acid increased the release of somatostatin, whereas E, and F,a prostaglandins had no effect. Our results show a close correlation between arachidonic acid release and somatostatin secretion, the two parameters we investigated.The transduction mechanisms of hypothalamic somatostatin (SRIF) release are not yet fully understood. We previously found that in hypothalamic cells in primary culture adenylate cyclase stimulation is not involved in SRIF release (I). Conversely, we showed that Ca" ionophores (A23187 or ionomycin) and K + depolarization (1, 2) stimulate SRIF secretion and that these effects depend on the presence of C a Z + in the extracellular media, but this process did not seem to be associated with the activation of a Ca2+-calmodulin kinase system ( 2 ) . More recently, we reported that glutamate strongly stimulates SRIF secretion in vitro (3) through N-methyl-D-aspartate (NMDA) receptor sites. Indeed, activation of NMDA receptors is known to greatly increase C a Z + entry into the cells through the NMDA receptor channel complex (4). These results suggest that SRIF release could be triggered by any type of C a 2 + entry into the cells. In view of recent data showing that NMDA receptors activate the arachidonic acid cascade system in striatal neurons (5) and cerebellar granule cells (6), presumably by activation of phospholipase A, ( 5 , 7), we investigated whether arachidonic acid release ~ .._______.could be modified after NMDA receptor activation in hypothalamic neurons in primary culture. We also examined whether pharmacological manipulation of phospholipase A, could modify SRIF release and studied the effect of exogeno...

GABA_A and NMDA Receptor Activation Controls Somatostatin Messenger RNA Expression in Primary Cultures of Hypothalamic Neurons

“…A nonspecific effect of MK-801 on DNA [28] and protein synthesis [29], as suggested in other systems, seems here unlikely since we previously measured total DNA and somatostatin content after a 24-hour treatment with MK-801 and found no effect of that drug on the two parame ters [14]. We also reported endogenous release of gluta mate which is strongly blocked by tetrodotoxin [11,13,30] and, more specifically, by CGS 19755, a NMDA receptor antagonist [13]. In addition, tonic activation of NMDA receptors by endogenous glutamate release was also shown in hippocampal slices [31 ].…”

Boeravinone B Protects Brain against Cerebral Ichemia Reperfusion Injury in Rats: Possible Role of Anti-inflammatory and Antioxidant

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