Satoshi Ohkura

Physiology of Reproduction

Publisher Summary The rat is the most popular experiment animal for studying reproductive physiology. Investigations have found that they have much shorter reproductive cycles compared with larger animals. Stages such as puberty, the estrous cycle, pregnancy, and lactation are all much shorter. In addition, rats display various types of estrous cycle, such as estrous cycle with luteal phase or spontaneous ovulation, when they receive certain stimuli. Rat sexual behavior also plays a significant role in the field of research. In order to observe sexual behavior in rats, a male rat that has already been trained to mate with normal females several times is placed in a test cage under a red dim light, e.g. a 60 x 50 x 40 cm plastic cage with bedding at the bottom to facilitate activity. After several minutes, a steroid-induced estrous female is introduced. The male starts an investigation and genital sniffing if he realizes that the female is in estrus. The male then shows mounting behavior with a pelvic thrust. After ejaculation, the male suddenly quits frequent pelvic thrusts and leaves the female. The female shows the lordosis behavior in response to the male mounting her.

Neural Control of Maternal Behaviour and Olfactory Recognition of Offspring

In terms of reproductive success the quality and duration of maternal care exhibited by any particular species is of paramount importance, and yet compared with the amount of research studying the control of reproductive cycles, sexual behaviour, and fertility, it has historically received considerably less attention. However, we are now beginning to understand how the brain is organised to mediate this complex behaviour and how its expression is orchestrated by different hormonal and neurochemical factors. This review summarises a series of neuroanatomical, electrophysiological, in vivo sampling and behavioural neuropharmacological experiments carried out in sheep. These have attempted to define the neural circuitry and hormonal neurotransmitter systems involved both in the control of maternal behaviour per se and in the selective olfactory recognition of lambs, which is the basis of an exclusive emotional bond between mother and offspring.

The Role of Oxytocin Release in the Paraventricular Nucleus in the Control of Maternal Behaviour in the Sheep

Oxytocin (OT) release within the brain is thought to play a major role in inducing maternal behaviour in a number of mammalian species but little is known about the sites of release which are important in this respect. We have investigated whether the paraventricular nucleus of the hypothalamus (PVN) is a site of OT action on maternal behaviour in the sheep. In vivo microdialysis and retrodialysis was used to determine whether OT is released in the region of the PVN during the post‐partum induction of maternal behaviour and if its release at this site can stimulate maternal behaviour in non‐pregnant animals. In vivo sampling showed that OT concentrations increased significantly in the region of PVN at birth. When OT was retrodialysed bilaterally into the PVN (1 or 10 μM) of multiparous ewes treated with progesterone and oestradiol to stimulate lactation, maternal behaviour was induced in a significant number of animals (1 μM, 6/8 and 10 μM, 5/8) compared with controls (0/8 ewes). Similar infusions of the ring structure of OT, tocinoic acid (TOC—10 μM), also induced maternal behaviour in a significant proportion of animals (5/6 ewes) as did intracerebroventricular (ICV) OT (6/8 ewes) and artificial stimulation of the vagina and cervix (VCS, 8/9 ewes). On the other hand, vasopressin (AVP) 1 μM did not induce maternal behaviour in any ewes and a 10 μM dose only induced it in 2/8 animals. The neurochemical changes accompanying the above treatments were also investigated. Noradrenaline concentrations increased in the PVN after the retrodialysis administration of OT 1 μM and 10 μM, TOC 10 μM and AVP 1 μM, OT ICV and VCS. Dopamine concentrations were also increased by OT 10 μM, TOC 10 μM, AVP 1 μM and OT ICV. Aspartate and glutamate concentrations were significantly reduced by retrodialysis infusions of OT 1 μM and AVP 1 and 10 μM but not by any other treatment. Finally, the retrodialysis infusions of OT and TOC, as well as ICV OT, significantly increased plasma OT release whereas AVP infusions did not. These results provide evidence that OT is released in the PVN during parturition and is important for the induction of maternal behaviour. It seems probable that OT release at this site has a positive feedback effect on both parvocellular and magnocellular OT neurones to facilitate co‐ordinated OT release both in central OT terminal regions (to facilitate maternal behaviour) and peripherally into the blood (to facilitate uterine contractions/milk let down). The potential functional roles for the actions of OT on monoamine and amino acid transmitter release in the PVN are discussed.

Intracerebroventricular Administration of Melanin-Concentrating Hormone Suppresses Pulsatile Luteinizing Hormone Release in the Female Rat

Melanin‐concentrating hormone (MCH) has been reported to be involved in the regulation of feeding behaviour in rats and mice. Because many neuropeptides that influence ingestive behaviour also regulate reproductive function, the present study was designed to determine if central administration of MCH changes pulsatile secretion of luteinizing hormone (LH) in the rats. Wistar‐Imamichi strain female rats were ovariectomized and implanted with oestradiol to produce a moderate inhibitory feedback effect on LH release. The effects of i.c.v. injections of MCH on LH release were examined in freely moving animals. Blood samples were collected every 6 min for 3 h through an indwelling cannula. After 1 h of sampling, MCH (0.1, 1 or 10 μg/animal) or vehicle (saline) was injected into the third cerebroventricle. Because MCH is also reported to affect the hypothalamo‐pituitary‐adrenal (HPA) axis, which in turn, can influence reproductive function, plasma corticosterone concentrations were determined in the same animals at 30‐min intervals during the first and last hours and every 12 min during the second hour of the 3‐h sampling period. When expressed as per cent changes, mean plasma LH concentrations after MCH administration were significantly lower in the animals injected with all doses of the peptide compared with vehicle‐treated animals; LH pulse frequency was significantly lowered by 1 μg of MCH. Per cent changes in mean plasma corticosterone levels were not significantly affected by MCH administration. These results in oestradiol‐treated ovariectomized rats indicate that central MCH is capable of inhibiting pulsatile LH secretion. We have previously shown that 48‐h fasting suppresses pulsatile LH release in the presence of oestrogen. Take together, these results raise the possibility that MCH could play a role in mediating the suppression of LH secretion during periods of reduced nutrition.

Adolescent Growth and Development in Japanese Macaques (Macaca fuscata) : Punctuated Adolescent Growth Spurt by Season

Adolescent growth of Japanese macaques (Macaca fuscata) was studied. Their growth is composed both of a seasonal cycle of acceleration and deceleration and of linear increases. There is a major growth spurt in linear dimension and body weight at the beginning of the breeding season of the third and fourth year of life in females and males respectively, when they mature reproductively. They show additional accelerated growth in the following year(s). These growth spurts, in total, are considered to correspond with the adolescent growth spurt in humans. Adolescent growth of Japanese macaques is characterized by a punctuation by slower growth and a later start, which is considered to be the product of adaptation to a strongly seasonal environment.

Biphasic role of dopamine on female sexual behaviour via D2 receptors in the mediobasal hypothalamus.

Dopamine has been implicated in the control of sexual behaviour, but its role seems quite complex and controversial. The aim of the present experiments was to investigate the effects of dopamine (DA) acting on D2 receptors in the mediobasal hypothalamus (MBH) on sexual behaviour in female sheep. To achieve this, the D2 agonist, quinpirole, was administered bilaterally via microdialysis probes into the MBH of ovariectomized ewes either before or after oestradiol (E2) administration. Quinpirole (100 ng/ml) infused for 6 h just before E2 hastened the onset of oestrus behaviour and the luteinizing hormone surge, whereas the same treatment given 6-12 h or 18-21 h after E2 decreased the intensity of sexual receptivity without affecting LH or prolactin secretion. We then tested the hypothesis that E2 stimulates the onset of oestrus partly by decreasing DA activation of D2 receptors. In this case the D2 antagonists pimozide or spiperone (100 ng/ml) were infused into the MBH via microdialysis probes for 11 h in the absence of E2 administration. A significant number of ewes showed induction of receptivity with both antagonists, although its intensity was significantly lower than that induced by E2. These treatments generally did not significantly alter extracellular concentrations of monoamines or aminoacids although quinpirole modulated the ability of sexual interactions to increase noradrenaline release. These experiments show that DA acts via D2 receptors in the MBH to control female sexual behaviour in a biphasic manner: the onset of sexual motivation and receptivity requiring an initial increase in activation followed by a decrease. This dual action could explain some of the controversies concerning DA action on sexual behaviour.

Circadian rhythm in circulating CD16-positive natural killer (NK) cells in macaque monkeys, implication of plasma cortisol levels.

The daily change in both percentage and absolute number of circulating major lymphocyte subset was determined with young Japanese monkeys and rhesus monkeys. The blood sample was collected at four hour-intervals beginning at 16:00 for 24 hours under the condition of applying tethering system by which blood samples could be collected without restraint. During the dark period (from 20:00 to 08:00), the number of peripheral lymphocytes increased and that of granulocytes decreased, resulting in no significant change in the number of total peripheral white blood cells. The absolute number of CD4 + T, CD8 + T, and CD20 + B cells showed the significant daily change similar to that in number of peripheral lymphocytes, indicating no proportional change in these subsets. The typical proportional change was observed in CD16 + natural killer (NK) cells and the percentage of CD16 + cells decreased during dark period (from 20:00 to 04:00) and increased in the morning (from 08:00 to 12:00). The NK activity determined by killing K562 target cells showed the same changing pattern as that of percentage in CD16+ NK cells. The changing pattern of both percentage and activity of NK cells was consistent with that of plasma cortisol levels. In addition, the intravenous injection of 300 µg/kg of cortisol induced increase in plasma cortisol levels and decrease in percentage of CD16 + NK cells during the first 60 min after cortisol injection. These results strongly suggest that the levels of peripheral functional CD16 + NK cells might be directly regulated by plasma cortisol level in macaque monkeys.

Chapter 20 – Endocrinology

Publisher Summary The rat is a good model for investigating the endocrine system, because the endocrine organs are of a reasonable size, being neither too small nor too big. This enables scientists to remove the organs or replace test substances easily at the laboratory level. Many different kinds of hormones have been isolated and identified in the rat, including hormones first identified in other animal species. This chapter reviews the functions of each endocrine organ, particularly their major hormones and the rat. Hypothalamic neuroendocrine cells of rats synthesize hypothalamic hormones that stimulate or inhibit the secretion of hormones from the anterior pituitary gland. These releasing/inhibiting hormones are discharged from the neuronal terminals located in the median eminence and diffuse into the capillaries where they are carried to the anterior pituitary gland via the pituitary portal system to regulate synthesis and secretion of anterior pituitary hormones. The gonads synthesize and secrete sex steroid hormones such as androgens, estrogens, and progestins. These gonadal steroids have a variety of actions, including stimulation of gamete production, development of reproductive organs, and expression of secondary sex characters and sexual behavior. Atrial natriuretic peptide (ANP) is known to act at the level of the kidney to increase sodium excretion into the urine. ANP also inhibits aldosterone release from the adrenal cortex. The thyroid gland produces two separate thyroid hormones called thyroxine/tetraiodothyronine (T4) and triiodothyronine (T3) under the control of thyroid-stimulating hormone. Parathyroid gland synthesizes and releases parathyroid hormone (PTH) from chief cells. PTH causes calcium mobilization from bones and enhances calcium reabsorption by the kidneys;these actions cause an elevation in plasma calcium concentration.