Growth-associated protein-43 messenger ribonucleic acid expression in gonadotropin-releasing hormone neurons during the rat estrous cycle

We have shown previously at the ultrastructural level that morphological ch anges occur in the external zone of the median eminence allowing certain Gn RH nerve terminals to contact the pericapillary space on the day of proestr us. The present study was designed to determine whether the intrinsic deter minant of neuronal outgrowth, growth-associated protein-43 (GAP-43), was ex pressed in GnRH neurons of adult female rats, and whether its expression va ried throughout the estrous cycle. To accomplish this, we perfusion-fixed g roups of adult female rats at 0800 and 1600 h on diestrous day 2 (diestrous II), at 0800 h and 1600 h on proestrus, and at 0800 and 1600 h on estrus ( n = 4 rats/group) and used double labeling in situ hybridization and quanti fication to compare the levels of GAP-43 messenger RNA (mRNA) in cells coex pressing GnRH mRNA. GnRH mRNA was detected with an antisense complementary RNA (cRNA) probe labeled with the hapten digoxigenin, whereas the GAP-43 cR NA probe was labeled with S-35 and detected by autoradiography. In addition , GAP-43 protein was identified with immunohistochemistry in the median emi nence. The results show that many GnRH neurons ex-pressed GAP-43 mRNA and t hat: GAP-43 protein was present in many GnRH axon terminals in the outer la yer of the median eminence. The number of GnRH neurons expressing GAP-43 mR NA was significantly higher on proestrus (64 +/- 5%) than on diestrous II ( 40 +/- 2%; P < 0.001) or on estrus (45 +/- 8%; P < 0.05), and the GAP-43 mR NA levels in GnRH neurons also varied as a function of time of death during the estrous cycle. The GAP-43 mRNA levels in GnRH neurons were higher on p roestrus and estrus than on diestrous II (P < 0.05). These data show that 1 ) GAP-43 is expressed in adult GnRH neurons; 2) GAP-43 mRNA expression in G nRH neurons fluctuates during the estrous cycle; and 3) GAP-43 mRNA content in GnRH neurons is highest on the day of proestrus, before and during the onset of the LH surge. These observations suggest that the increased GAP-43 mRNA expression in GnRH neurons on the day of proestrus could promote the outgrowth of GnRH axon terminals to establish direct neurovascular contacts in the external zone of the median eminence and thus facilitate GnRH relea se into the pituitary portal blood.