A NOVEL NEURAL PATHWAY CONTROLS TESTOSTERONE RELEASE

我们最近发现了一个神经脑睾丸通路的存在，干扰Leydig细胞功能独立的垂体。 具体地说，我们发现在侧脑室（icv）注射促肾上腺皮质激素释放因子（CRF）或异丙肾上腺素（ISO）后5分钟内，人绒毛膜促性腺激素（hCG）对睾酮（T）释放的刺激作用被阻断。 在hCG之前15分钟注射全身酒精产生类似的效果。 icv CRF或ISO或酒精阻断T反应的能力既不被模仿，也不被GnRH拮抗剂iv预处理逆转，表明它们的抑制作用不是由于LH水平低。 相反，icv注射肾上腺素能拮抗剂部分逆转icv ISO，icv CRF或全身酒精的作用。这些结果支持存在通过肾上腺素能机制快速抑制间质细胞功能的神经通路。 目前，这一途径的神经形态学证据缺失。 在特定目标1下，我们将使用病毒跨神经元标记方法来鉴定中枢神经系统（CNS）中参与该途径的位点。 这种强大的神经解剖工具包括将伪狂犬病病毒（PRV）注射到睾丸中。 在复制之后，PRV以逆行的方式被运输到支配性腺的一级神经元的胞体，然后随着感染进行到进一步的突触连接的细胞体而被运输到二级、三级和四级神经元。 通过使用多克隆抗体的免疫细胞化学鉴定病毒标记的神经元。 标记的特异性将通过精索去神经支配和/或脊髓T1切片的能力来确定，以消除更高结构中的PRV标记。 在我们确定了属于拟议通路的大脑区域后，我们将进行功能实验以确定其生理重要性。 在特定目标2下，我们将在选定的大脑区域微灌注CRF或ISO。 我们预计，通过激活拟议的抑制途径，这些治疗将减少hCG诱导的T分泌，与输注溶媒的动物中获得的结果相比。 在特定目标3下，我们将在T1水平切断脊髓或损伤被认为参与拟定途径的特定下丘脑区域，以确定这些程序是否阻断icv CRF或全身酒精对hCG诱导的T分泌的抑制作用。 这些实验将提供第一个形态和功能的证据，睾丸和中枢神经系统之间的多突触神经通路的存在，这是酒精的影响。

We recently uncovered the existence of a neural brain-testicular pathway that interferes with Leydig cell function independently of the pituitary. Specifically, we showed that within 5 min of their intracerebroventricular (icv) injection, corticotropin- releasing factor (CRF) or isoproterenol (ISO) blocked the stimulatory effect of human chorionic gonadotropin (hCG) on testosterone (T) release. Systemic alcohol injected 15 min prior to hCG exerted a similar effect. The ability of icv CRF or ISO, or of alcohol, to block the T response was neither mimicked nor reversed by iv pretreatment with a GnRH antagonist, indicating that their inhibitory effect was not due to low LH levels. In contrast, the icv injection of an adrenergic antagonist partially reversed the effect of icv ISO, icv CRF or systemic alcohol. These results support the existence of a neural pathway that rapidly inhibits Leydig cell function through an adrenergic mechanism. At present, neuromorphological evidence for this pathway is missing. Under Specific Aim 1, we will use a viral transneuronal labeling method to identify sites in the central nervous system (CNS) that are involved in this pathway. This powerful neuroanatomical tool consists of the injection of pseudorabies virus (PRV) into the testis. Following replication, PRV is transported in a retrograde fashion to the perikarya of first-order neurons innervating the gonad, then to second-, third- and fourth-order neurons as the infection proceeds to further synaptically linked cell bodies. Virus-labeled neurons are identified by means of immunocytochemistry using polyclonal antibodies. Specificity of labeling will be determined by the ability of spermatic denervation and/or spinal cord T1 section, to eliminate PRV labeling in higher structures. After we have identified brain areas that belong to the proposed pathway, we will conduct functional experiments to determine its physiological importance. Under Specific Aim 2, we will microinfuse CRF or ISO in selected brain areas. We anticipate that by activating the proposed inhibitory pathway, these treatments will decrease hCG-induced T secretion, compared to results obtained in animals infused with the vehicle. Under Specific Aim 3, we will sever the spinal cord at the T1 level or lesion specific hypothalamic areas thought to be involved in the proposed pathway, to determine whether these procedures block the inhibitory effect of icv CRF or systemic alcohol on hCG-induced T secretion. These experiments will provide the first morphological and functional evidence of the existence of a multisynaptic neural pathway between the testes and the CNS, that is influenced by alcohol.