INTRACELLULAR CA2+ IN LHRH NEURONS FROM RHESUS EMBRYONIC OLFACTORY PLACODE

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OBJECTIVE To understand the mechanism of pulsatile LHRH release, and whether cultured LHRH neurons exhibit spontaneous intracellular Ca2+ ([Ca2+]i) signaling. The olfactory placode and the ventral migratory pathway of LHRH neurons from rhesus monkey embryos at E35-37 were dissected out and cultured on glass coverslips. Two to 5 weeks later cultured cells were labeled with the calcium dye, fura-2, and examined for [Ca2+]i signaling by recording changes in [Ca2+]i every 10 s for 30-175 min. Cells were fixed and immunostained for LHRH and neuron specific enolase. In 20 cultures, 572 LHRH-positive cells exhibited [Ca2+]i oscillations at an interpulse interval (IPI) of 8.2(0.7 min, and a duration of 88.8(2.9 s. LHRH-negative neurons in culture exhibited only occasional [Ca2+]i oscillations. In 17 of 20 cultures with LHRH-positive cells, [Ca2+]i oscillations occurred synchronously in 50-100% of the individual cells, whereas [Ca2+]i oscillations in cells in the remaining 3 cultures did not synchronize. Strikingly, in 12 of 17 cultures the synchronization of [Ca2+]i oscillations repeatedly occurred in complete unison at 52.8(3.0 min intervals, which is similar to the period observed for LHRH release, whereas in 5 of 17 cultures the less tight synchronization of [Ca2+]i oscillations repeatedly occurred at 23.4(4.6 min intervals. IPI of [Ca2+]i oscillations in cells with tight synchronization and less tight synchronization did not differ from IPI in cells without synchronization. The results indicate that LHRH neurons derived from the monkey olfactory placode possess an endogenous mechanism for synchronization of [Ca2+]i oscillations. FUTURE DIRECTIONS We will determine if synchronization of [Ca2+]i oscillations is manifestation of periodical LHRH neurosecretion. KEY WORDS intracellular Ca2+ signaling, intracellular Ca2+ oscillations, synchronization, LHRH neurons, olfactory placode. FUNDING NIH HD15433 & RR00167 PUBLICATIONS Terasawa, E. Cellular mechanism of pulsatile LHRH release. Gen. Comp. Endocrinol. 112 283-295, 1998. (J)