• Neurotransmitter Gases as Modulators of GnRH and the Preovulatory LH Surge

      Lamar, Charisee; Institute of Molecular Medicine and Genetics (1998-04)
      Until recently nitric oxide (NO) and carbon monoxide (CO) were viewed only as toxic substances. However, there has been a substantial amount of evidence in the past decade that has redefined these gaseous molecules as physiological messengers. Along these lines, NO and CO are now recognized as modulators of immunological defense, vasodilation, endocrine signaling, and neurotransmission (1-8). As neurotransmitters, NO and CO are unique when compared to classical neurotransmitters. For instance, unlike all other classical neurotransmitters NO and CO are, 1) lipophilic gases with short half lives, 2) not stored in synaptic vesicles, 3) their effects are not mediated through classic receptor proteins - rather their effects result from NO and CO binding to the heme moiety of heme-proteins such as guanylate cyclase and cyclooxygenase, and 4) their effects are terminated by diffusion from target tissues ( 1,2,5,7,9). Production of NO and CO relies on the activity of the enzymes nitric oxide synthase (NOS) and heme oxygenase (HO), respectively. NOS uses the substrate Larginine to generate NO (2, 10), while HO uses the substrate heme to generate CO (9,11-16). NOS exists as three isoforms, macrophage NOS (mNOS), endothelial NOS (eNOS), and neuronal NOS (nNOS) (17-21). HO also exists as three isoforms, heme oxygenase-1 (HO-1), heme oxygenase-2 (HO-2), and heme oxygenase-3 (HO-3) HO-1 is inducible, while HO-2 and HO-3 are constitutive enzymes; however, HO-3 is currently viewed as a poor heme catalyst (12,13,22). There is a growing body of evidence that suggests that NO and CO regulate hypothalamic function. For example, recent studies have shown that the hypothalamus produces significant quantities of NO, primarily through the actions of nNOS (23,24). Likewise, the hypothalamus possesses one of the largest production rates of CO in the brain (25,26). That NO and CO can regulate neurohormone release from the hypothalamus is evidenced by findings demonstrating that NO and CO regulate corticotropin-releasing hormone (CRH) (27-33), vasopressin (29-31,34,35), and oxytocin secretion (29,36). With respect to reproductive function, numerous studies (23,37-45) have provided evidence for a significant role of NO in the control of the hypothalamic releasing factor, gonadotropin-releasing hormone (GnRH). For instance, it has been demonstrated (37,46,47) that NO neurons are located in close proximity to GnRH generating cells bodies in the hypothalamus and thus strategically located to exert regulatory effects over GnRH neurons. That NO can exert regulatory effects on GnRH neurons was demonstrated in studies where exogenously applied NO markedly stimulated GnRH release from male hypothalamic fragments (1,48,49) and immortalized GnRH (GT1-7) cells in vitro (1,48). A physiological role for NO in the steroid-induced luteinizing hormone (LH) surge has been suggested based on studies in which the LH surge was attenuated by the administration of NOS inhibitors (43,44) or NOS antisense oligonucleotides (38) No studies, however, have reported nNOS mRNA, protein, or NOS activity levels in the hypothalamus during the LH surge so as to verify that an increased NO tone actually occurs during this critical time. To address this deficit, Aim 1 of this study was designed to examine NO tone on proestrus in the cycling female rat-the day the natural preovulatory LH surge occurs. Since heme oxygenase, the enzyme that produces CO from heme molecule metabolism, is located in the hypothalamus (2,11,12,14,15), and CO production in the hypothalamus is one of the highest in the brain (25,26), it is conceivable that CO could play a role in regulating GnRH secretion. This possibility has not been investigated and thus studies on this issue appear warranted. Therefore, Aims 2-4 of this study were designed to assess the possible role of CO in the control of GnRH and LH secretion.