Atsuhiko Chiba

Effects of sexual experience on conspecific odor preference and male odor-induced activation of the vomeronasal projection pathway and the nucleus accumbens in female rats.

In the present study in estrogen-progesterone primed ovariectomized female rats, we examined the expression of a preference for male odors and male odor-induced Fos immunoreactivity throughout the vomeronasal projection pathway and the nucleus accumbens (NAcc), using both sexually experienced and sexually naive subjects. Female rats significantly preferred airborne odors and soiled bedding from sexually active males over those from estrous females, irrespective of the presence or absence of prior sexual experience. On the other hand, the brain regions in which exposure to male-soiled bedding significantly increased Fos expression were different between sexually experienced and sexually naive subjects. Significant increment of Fos expression in the posterior-dorsal medial amygdala (MePD) and the bed nucleus of stria terminalis (BNST) in forebrain, as well as the accessory olfactory bulb, was observed in both groups of subjects. Fos expression in the anterior-dorsal medial amygdala (MeAD), the medial preoptic area (mPOA) and the NAcc core, however, was significantly increased only in the sexually experienced subjects. These results suggested that male odor-induced activations of the MePD and/or the BNST, but not of the MeAD, the mPOA and the NAcc core, are required for the expression of a male-directed odor preference in female rats.

On the Relationship between Noradrenergic Stimulatory and GABAergic Inhibitory Systems in the Control of Luteinizing Hormone Secretion in Female Rats

The relationship between noradrenergic (NA) stimulatory and gamma-aminobutyric acid (GABA)-mediated inhibitory systems in the control of luteinizing hormone (LH) secretion was examined in ovariectomized rats. Stimulation of the GABAA receptor by an intraventricular (i.c.v.) administration of a GABAA agonist, muscimol, significantly attenuated the LH secretory response to the subsequent i.c.v. injection of NA in estrogen-primed rats. On the other hand, blockade of alpha-adrenergic receptors by phenoxybenzamine did not interfere with the stimulatory effect of an i.c.v. injection of a GABAA antagonist bicuculline. In a different series of experiments, ovariectomized animals had been treated with i.c.v. injections of 6-hydroxydopamine (6-OHDA) or its vehicle. An i.c.v. injection of muscimol or phentolamine significantly inhibited the estrogen-induced surge of LH secretion in vehicle-treated rats. Muscimol also inhibited the existing pulsatile LH secretion in vehicle-treated, estrogen-unprimed animals. In animals in which hypothalamic NA terminals were presumed to be destroyed by 6-OHDA, the inhibitory effect of phentolamine was significantly diminished while that of muscimol was unaltered. These results permit the following conclusions: (1) the central GABAergic system inhibits LH secretion via GABAA receptors; (2) this inhibitory GABA system operates without mediation by the NA system, and (3) the GABAergic non-NA-mediated system can affect physiological patterns of pituitary LH secretion in female rats.

Acute Immobilization Stress and Intraventricular Injection of CRF Suppress Naloxone‐Induced LH Release in Ovariectomized Estrogen‐Primed Rats

The present study was undertaken to evaluate the role and possible interaction of the endogenous opioid peptide (EOP) and corticotropin‐releasing factor (CRF) in the acute stress‐induced suppression of gonadotropin secretion in ovariectomized estrogen‐primed rats. An intravenous (i.v.) injection of naloxone (10 or 20 mg/kg), an EOP antagonist, significantly elevated serum luteinizing hormone (LH) levels within 10 min in non‐stressed animals. The naloxone‐induced LH release was completely eliminated when tested 30 min after the onset of acute immobilization. In a subsequent study, it was found that suppression of the naloxone‐induced LH release occurred as early as 5 min after the stress onset, and was still evident 60 min after the end of a 30‐min period of immobilization. The effect of naloxone was restored 3 h after liberation of the animal from the 30‐min immobilization. An intraventricular (i.c.v.) injection of CRF (1 or 5 μg) also significantly suppressed, in a dose‐related manner, the effect of a subsequent i.v. injection of naloxone. However, an i.c.v. injection of α‐helical CRF(9‐41) (25 or 50 μg), a CRF antagonist, prior to immobilization, could not interfere with the suppressive effect of stress on naloxone‐induced LH release. These results suggest that both acute immobilization stress and CRF can inhibit the LH secretory activity without mediation by EOP neurons. However, the stress‐related suppression may involve non‐CRF mechanism(s).

Permissive role of corticotropin-releasing factor in the acute stress-induced prolactin release in female rats

The present study examined the role of corticotropin-releasing factor (CRF) in the acute stress-induced release of prolactin (PRL) in ovariectomized estrogen-primed female rats. Acute immobilization stress induced a marked increase in serum PRL levels in animals treated with saline intraventricularly (i.c.v.). However, a prior icv injection of alpha-helical CRF(9-41), a CRF antagonist, completely eliminated the immobilization-induced PRL release in the majority of animals, providing evidence for involvement of CRF in the acute stress-induced PRL release. On the other hand, an i.c.v. injection of CRF did not affect basal PRL release at any dose in non-stressed animals, suggesting that the peptide plays a permissive role which enables other undefined stress mediator(s) to stimulate PRL release.

Effects of pinealectomy and melatonin on the timing of the proestrous luteinizing hormone surge in the rat.

Effects of pinealectomy and melatonin replacement on the timing of the preovulatory luteinizing hormone (LH) surge were examined in female rats housed under a light-dark cycle (lights on 06:30-18:30 h). Animals were pinealectomized or sham-operated 21-28 days before bleeding. They were sequentially bled in the afternoon of proestrus through the indwelling cardiac cannula without anesthesia. Proestrous LH surges were observed in all pinealectomized rats, but the onset times of the LH surge in these animals showed a significantly greater variance than those in sham-operated controls. A single melatonin injection at 18:00 or 21:00, shortly before or after the light-dark transition, on the day before proestrus (diestrus II) reduced the variance in the LH surge onset in pinealectomized rats in a dose-dependent manner. In contrast, melatonin injected at 14:00 or 23:00 on diestrus II or at 04:00 on proestrus was ineffective. These results showed that the pineal gland is involved in controlling the timing of the proestrous LH surge in the rat. The pineal signal may be transmitted by melatonin, a major product of the gland.

Effects of androgens and estrogens on crowings and distress callings in male Japanese quail, Coturnix japonica.

In male Japanese quail, crowing behavior is considered to be strictly androgen-dependent. It was previously shown that in chicks, treatment with either testosterone or 5alpha-dihydrotestosterone (5alpha-DHT; a non-aromatizable androgen) induced crowing with motivation for distress calling in acutely isolated conditions. Many studies, however, have shown that the potencies of testosterone and 5alpha-DHT in activating crowing in castrated males are different. To clarify the effects of androgenic and estrogenic actions on the production of crows and distress calls, we injected quail daily from 11 to 42 days after hatching (Day 11 to 42) with testosterone propionate (TP), 5alpha-DHT, estradiol benzoate (EB) or vehicle and examined their calling behaviors both in a recording chamber (acutely isolated conditions) and in their home-cages (well-acclimated conditions). Both TP- and 5alpha-DHT-treated birds began to crow by Day 13 when isolated in the recording chamber. The TP-treated birds, however, crowed less frequently than 5alpha-DHT-treated ones. This, combined with the observations that distress calling was strongly inhibited in EB-treated birds, suggests that estrogen converted from testosterone may inhibit the motivation for distress calling. On the other hand, after chronic treatment of TP, but not of 5alpha-DHT, birds began to crow intensely in their home-cages earlier than vehicle treated controls, suggesting that estrogen is needed to initiate crowing behavior in sexually active males. Taken together, it is suggested that estrogenic actions affect the motivation underlying vocal behaviors, while the androgenic action is indispensable in generating crowing.

Androgen receptor blockade in the posterodorsal medial amygdala impairs sexual odor preference in male rats.

The present study was designed to investigate the role of androgen in the medial amygdala (MeA) in the expression of sexual odor preference in male rats. Gonadally intact, sexually experienced male rats received bilateral administration of flutamide, an androgen receptor (AR) blocker, aimed at either the posterior dorsal part (MePD) or the anterior dorsal part (MeAD) of the MeA through inner cannulae inserted into the implanted guide cannulae. Prior to flutamide administration, all subjects spent longer sniffing volatile odors from an estrous female than those from a sexually active male. Experiment 1 demonstrated that the preference for the female odors over the male odors was eliminated during flutamide administration into the MePD, but not into either the MeAD or outside MePD/MeAD. This elimination of the female-directed odor preference resulted from increase of time sniffing the male odors rather than decrease of time sniffing the estrous odors. In Experiment 2, odor discrimination tests confirmed that the flutamide administration into the MePD did not induce impairment in the ability of the subjects to discriminate the estrous odors from the male odors. These results demonstrated that activation of AR in the MePD plays a critical role in the expression of the preference for estrous odors over male odors. AR blockade, however, seemed to induce a preference for male odors rather than reduce the existing preference for estrous odors, suggesting a complicated regulation of sexual odor preference by sex steroids.

Acute Stress Suppresses the N-Methyl-D-Aspartate-Induced Luteinizing Hormone Release in the Ovariectomized Estrogen-Primed Rat

The effects of N-methyl-D-aspartate (NMDA) and luteinizing hormone releasing hormone (LH-RH) on luteinizing hormone (LH) secretion were examined in ovariectomized estrogen-primed rats under nonstressed and acutely stressed conditions. The basal LH levels were significantly elevated 15 min after the onset of acute immobilization stress, but were not altered in emotionally stressed or nonstressed rats. Intravenous injections of 10 and 40 mg/kg NMDA significantly elevated serum LH levels by 161 and 212%, respectively, from baseline within 10 min in nonstressed animals. However, the NMDA-induced LH release was significantly reduced when tested 30 min after the onset of acute immobilization stress. Acute emotional stress, which did not affect the baseline LH, also suppressed the LH release response to NMDA, suggesting that the reduced LH responses to NMDA in stressed animals was not due to the elevated baseline level. Pituitary LH release responses to LH-RH were not affected by acute immobilization. We conclude from these results: (1) acute immobilization stress exerts both stimulatory and inhibitory effects on LH release, while acute emotional stress has only an inhibitory effect in estrogen-primed ovariectomized rats; (2) this inhibition occurs at the suprapituitary level, and (3) it involves a suppression of the responsiveness of the hypothalamic LH-RH neuronal system to the excitatory amino acid input.

The effects of pinealectomy and blinding on the circadian locomotor activity rhythm in the Japanese newt, Cynops pyrrhogaster

We examined the effects of pinealectomy and blinding (bilateral ocular enucleation) on the circadian locomotor activity rhythm in the Japanese newt, Cynops pyrrhogaster. The pinealectomized newts were entrained to a light-dark cycle of 12 h light and 12 h darkness. After transfer to constant darkness they showed residual rhythmicity for at least several days which was gradually disrupted in prolonged constant darkness. Blinded newts were also entrained to a 12 h light/12 h dark cycle. In subsequent constant darkness they showed free-running rhythms of locomotor activity. However, the freerunning periods noticeably increased compared with those observed in the previous period of constant darkness before blinding. In blinded newts entrained to the light/dark cycle the activity rhythms were gradually disrupted after pinealectomy even in the presence of the light/dark cycle. These results suggest that both the pineal and the eyes are involved in the newts circadian system, and also suggest that the pineal of the newt acts as an extraretinal photoreceptor which mediates the entrainment of the locomotor activity rhythm.

ENTRAINMENT OF THE CIRCADIAN LOCOMOTOR ACTIVITY RHYTHM IN THE JAPANESE NEWT BY MELATONIN INJECTIONS

We examined whether melatonin can act as a synchronizing agent within the circadian system of amphibians by testing the ability of melatonin injections to entrain the circadian locomotor activity rhythm of a newt (Cynops pyrrhogaster). Under constant darkness, all newts (13 cases) showing the free-running rhythms were subcutaneously injected with 10 μg melatonin at the same time every other day for at least 30 days. Subsequently, they were injected with vehicle (1% ethanolic saline) instead of melatonin for at least another 30 days. In 10 of the 13 newts, the locomotor activity rhythms could be entrained to a period of 24 h by melatonin injections but not by vehicle injections. During the entrained steady-state, the active phase of an activity-rest cycle preceded the time of melatonin injections as previously reported in other diurnal species. These results suggest that the endogenous circadian rhythm of melatonin concentration may be involved in synchronizing circadian oscillator(s) within the newts circadian system.