Evidence for a spontaneous nitric oxide release from the rat median eminence: Influence on gonadotropin-releasing hormone release

The involvement of nitric oxide (NO) as a gaseous neurotransmitter in the h ypothalamic control of pituitary LH secretion has been demonstrated. NO, as a diffusible signaling gas, has the ability to control and synchronize the activity of the neighboring cells. NO is secreted at the median eminence ( ME), the common termination field for the antehypophysiotropic neurons, und er the stimulation of other signaling substances. At the ME, NO stimulates GnRH release from neuroendocrine terminals. The present studies were undert aken to determine whether NO is secreted spontaneously from ME fragments ex vivo and whether its secretion is correlated to GnRH release. To accomplis h this, female rats were killed at different time points of the day and/or of the estrous cycle. The spontaneous NO release was monitored in real time , with an amperometric probe, during 4 periods of 30 min, from individual M E fragments (for each time point, n = 4). GnRH levels were measured in para llel for each incubation-period by RIA. The results revealed that NO was re leased in a pulsatile manner from female ME fragments and, unambiguously, t hat the amplitude of NO secretion varied markedly across the estrous cycle. Indeed, though the NO pulse period (32 +/- 1 min, n = 36) and duration (21 +/- 2 min, n = 36) did not vary significantly across the estrous cycle, th e amplitude of this secretion pulse was significantly higher on proestrus ( Pro; 39 +/- 3 nhl, n = 20), compared with diestrus (16 +/- 1 nM, n = 8) or estrus (23 +/- 3 nM, n = 8, P < 0.05). The GnRH levels in the incubation me dium were positively correlated to NO secretion across the estrous cycle (r = 0.86, P < 0.003, n = 9), confirming that NO and GnRH release are coupled . Furthermore, 5 x 10(-7) M L-N-5-(1-iminoethyl)ornithine (L-NIO), a NO syn thase inhibitor, succeeded in inhibiting the strong NO-GnRH secretory coupl ing and GnRH release on Pro. Because at this concentration, L-NIO selective ly inhibits endothelial NO synthase, the results further demonstrate that t he major source of NO involved in GnRH release at the ME is endothelial in origin. Additionally, the induction of a massive NO/GnRH release in Ig-day ovariectomized rat treated with estradiol benzoate strongly suggested that estradiol is participating in the stimulation of NO release activity betwee n diestrus II and Pro. The present study is the first demonstrating that ME can spontaneously release NO and that NO's rhythm of secretion varies mark edly across the estrous cycle. This pulsatile/cyclic ME NO release may cons titute the synchronizing link to anatomically scattered GnRH neurons.