Toru Ichimaru

Gonadotrophin‐Releasing Hormone Pulse Generator Activity in the Hypothalamus of the Goat

Pulsatile release of gonadotrophin‐releasing hormone (GnRH) is indispensable to maintain normal gonadotrophin secretion. The pulsatile secretion of GnRH is associated with synchronised electrical activity in the mediobasal hypothalamus (i.e. multiple unit activity; MUA), which is considered to reflect the rhythmic oscillations in the activity of the neuronal network that drives pulsatile GnRH secretion. However, the cellular source of this ultradian rhythm in GnRH activity is unknown. Direct input from kisspeptin neurones in the arcuate nucleus (ARC) to GnRH cell bodies in the medial preoptic area or their terminals in the median eminence could be the intrinsic source for driving the GnRH pulse generator. To determine whether kisspeptin signalling could be responsible for producing pulsatile GnRH secretion, we studied goats, measured plasma levels of luteinising hormone (LH) and recorded MUA in the posterior ARC, where the majority of kisspeptin neuronal cell bodies are located. Rhythmic volleys of MUA were found to be accompanied by LH pulses with regular intervals in the ARC, where kisspeptin neuronal cell bodies were found. Exogenous administration of kisspeptin stimulated a sustained increase in LH secretion, without influencing MUA, suggesting that the GnRH pulse generator, as reflected by MUA, originated from outside of the network of GnRH neurones, and could plausibly reflect the pacemaker activity of kisspeptin neurones, whose projections reach the median eminence where GnRH fibres project. These observations suggest that the kisspeptin neurones in the ARC may be the intrinsic source of the GnRH pulse generator.

Structure and chemical organization of the accessory olfactory bulb in the goat.

The structure and chemical composition of the accessory olfactory bulb (AOB) were examined in male and female goats. Sections were subjected to either Nissl staining, Klüver‐Barrera staining, lectin histochemistry, or immunohistochemistry for nitric oxide synthase (NOS), neuropeptide Y (NPY), tyrosine hydroxylase (TH), dopamine β‐hydroxylase (DBH), and glutamic acid decarboxylase (GAD). The goat AOB was divided into four layers: the vomeronasal nerve layer (VNL), glomerular layer (GL), mitral/tufted (M/T) cell layer (MTL), and granule cell layer (GRL). Quantitative and morphometric analyses indicated that a single AOB contained 5,000–8,000 putative M/T cells with no sex differences, whereas the AOB was slightly larger in males. Of the 21 lectins examined, 7 specifically bound to the VNL and GL, and 1 bound not only to the VNL, but also to the MTL and GRL. In either of these cases, no heterogeneity of lectin staining was observed in the rostrocaudal direction. NOS‐, TH‐, DBH‐, and GAD‐immunoreactivity (ir) were observed in the MTL and GRL, whereas NPY‐ir was present only in the GRL. In the GL, periglomerular cells with GAD‐ir were found in abundance, and a subset of periglomerular cells containing TH‐ir was also found. Double‐labeling immunohistochemistry revealed that virtually all periglomerular cells containing TH‐ir were colocalized with GAD‐ir. Anat Rec, 2007.

Gene Expression Profiles Linked to the Hormonal Induction of Male-Effect Pheromone Synthesis in Goats (Capra hircus)

Abstract The male effect is a well-known phenomenon in female sheep and goats whereby a pheromone-induced activation of reproductive function occurs. However, the molecule(s) involved in this phenomenon are unknown. We investigated gene expression profiles for the induction of male effect pheromone synthesis using a PCR-based cDNA subtraction strategy. We constructed two subtracted cDNA libraries using mRNA from the skin of the head or rump region of orchidectomized male goats with or without pheromone induction using testosterone or dihydrotestosterone (DHT). Both libraries were assumed to contain genes whose expression increases with pheromone induction. Clones (n = 480) from each library were sequenced and identified using BLAST to reveal 115 and 239 types of sequences in the libraries of the head and rump region, respectively. Among these, 12 genes were expressed in both libraries. We conducted real-time PCR to further analyze their expression using cDNA samples derived from pheromone-producing or nonproducing skin from the head of an ovariectomized female goat with or without DHT implantation, respectively. For nine genes, we observed significantly increased expression in samples following DHT implantation. Among these, stearoyl-CoA desaturase 1 (SCD1) and elongation of long chain fatty acids family member 5 (ELOVL5) genes showed more than 100-fold higher expression levels in pheromone-positive samples, suggesting that the products of these genes may be important in pheromone synthesis.

Central Cholecystokinin‐Octapeptide Accelerates the Activity of the Hypothalamic Gonadotropin‐Releasing Hormone Pulse Generator in Goats

To clarify central actions of cholecystokinin‐octapeptide (CCK‐8) on reproduction, effects of an intracerebroventricular (i.c.v.) administration of CCK‐8 on the activity of the gonadotropin‐releasing hormone (GnRH) pulse generator were examined in ovariectomized (OVX) goats in the absence or presence of oestradiol. Goats were chronically fitted with recording electrodes in the mediobasal hypothalamus, and electrophysiological manifestations of the GnRH pulse generator were monitored as characteristic increases in the multiple‐unit activity (MUA volleys). In OVX goats, a bolus i.c.v. injection of as little as 0.01 nmol of CCK‐8 induced a MUA volley with a short latency, which resulted in a significant decrease in the post‐treatment volley interval compared to that in the saline injected control. Administration of higher doses of CCK‐8 (0.1 and 2 nmol) did not further accelerate the occurrence of the MUA volley, but stimulatory effects were observed for a longer period than that after the 0.01 nmol injection. When goats were treated with oestradiol, while a bolus i.c.v. injection of 0.01 nmol CCK‐8 had no effect, an injection of 0.1 nmol of the peptide significantly decreased the post‐treatment volley interval. On continuous i.c.v. infusion of CCK‐8 at 3 nmol per 200 µl/h for 3 h, MUA volleys with shorter intervals than those in the control were successively induced without any apparent change in basal plasma luteinizing hormone levels in OVX goats. These results demonstrate that central CCK‐8 strongly accelerates the activity of the GnRH pulse generator in goats.

Effects of exposure to male goat hair extracts on luteinizing hormone secretion and neuronal activation in seasonally anestrous ewes.

ABSTRACT In sheep and goats, exposure of seasonally anestrous females to males or their fleece/hair activates the gonadotropin-releasing hormone (GnRH) pulse generator leading to pulsatile luteinizing hormone (LH) secretion. Pheromones emitted by sexually mature males are thought to play a prominent role in this male effect. In the present study, we first aimed to clarify whether the male goat pheromone is effective in ewes. Seasonally anestrous St. Croix ewes were exposed to hair extracts derived from either intact or castrated (control) male Shiba goats. The male goat-hair extract significantly increased LH secretion compared to the control, suggesting that an interspecies action of the male pheromone occurs between sheep and goats. Using the male goat-hair extract as the pheromone source, we then aimed to clarify the neural pathway involved in the signal transduction of the male pheromone. Ewes were exposed to either the goat-hair extract or the control and sacrificed 2 hr after the exposure. Expression of c-Fos, a marker of neuronal activation, was immunohistochemically examined. The male goat-hair extract significantly increased the c-Fos expression compared to the control in regions of the vomeronasal system, such as the accessory olfactory bulb and medial amygdala, and the arcuate nucleus. The main olfactory bulb did not exhibit any significant increase in the c-Fos expression by the male goat-hair extract. This result suggests that the neural signal of the male pheromone is conveyed to the GnRH pulse generator through the activated regions in ewes.