PARENT SESSION

(144) BETA-ENDORPHIN INHIBITS ACTION-POTENTIAL FIRING FROM NEURONS EXPRESSING A GONADOTROPIN RELEASING HORMONE (GnRH)-GREEN FLUORESCENT PROTEIN (GFP) TRANSGENE IN THE TERMINAL NERVE OF TRANSGENIC FISH (MEDAKA, Oryzias latipes).

Wayne, Nancy ¹, Kuwahara, Kenrick¹, Aida, Katsumi², Nagahama, Yoshitaka³, Okubo, Kataaki³, ¹ UCLA School of Medicine, Los Angeles, CA² The University of Tokyo, Tokyo, Japan³ National Institute for Basic Biology, Okazaki, Aichi, Japan

ABSTRACT- The electrical activity of GnRH neurons is thought to regulate secretion of GnRH, and thus be a critical determinant of the reproductive state of vertebrates. Factors that modulate GnRH secretion are likely doing so through changes in membrane excitation of the neurons. Activation of endogenous opioid receptors inhibits GnRH secretion, leading to a suppression of reproductive function. In this study, we tested the hypothesis that opiates inhibit the electrical activity of GnRH neurons. Experiments were performed on a newly developed line of transgenic medaka in which a salmon (s)GnRH-GFP transgene is expressed in the terminal nerve (TN) of the olfactory bulb. This population of sGnRH neurons has been shown to regulate reproductive behavior in male teleost fish. Expression of the transgene allows for identification of GnRH neurons for targeted electrophysiology. Whole-cell current clamp recordings were performed in the excised, intact brain of adult males with stable electrophysiological recordings routinely lasting over 60 min. Under control conditions, the TN-sGnRH-GFP neurons (n=13 cells from 9 fish) had a resting membrane potential of -48.4 ± 1.3 mV (mean ± sem) and were spontaneously active with a spike frequency of 1.35 ± 0.18 Hz. The action potentials exhibited an amplitude of 95.8 ± 3.2 mV, and a width of 3.05 ± 0.35 msec. Bath application of 1 microM beta-endorphin led to a gradual decline in the amplitude of the spikes compared to the baseline period, with a 12% decrease by 10 min (not significant), 26% decrease by 20 min (P < 0.05), and 57% decrease by 30 min of treatment (P < 0.01) (n=6 cells from 5 fish). In 2 of 6 cells, by 30 min of treatment the oscillation in membrane depolarization remained, but did not reach threshold to fire action potentials. There was a partial washout affect, with action-potential amplitude recovering to 75% of the baseline value. These findings support the hypothesis that endogenous opiates suppress GnRH secretion by inhibiting action-potential firing.

KEY WORDS: gonadotropin releasing hormone, electrophysiology, transgenic, neuroendocrine