Chandra Mohini Chaturvedi

An in situ hybridization and immunohistochemical study of vasotocin neurons in the hypothalamus of water-deprived chickens

The distribution of immunoreactive vasotocin (IR-AVT) and AVT mRNA in the hypothalamus of White Leghorn cocks was determined by immunohistochemistry and in situ hybridization, respectively. In control birds that were provided with water ad lib, AVT mRNA was distributed in the periventricular and lateral regions of the hypothalamus in clusters of neurons that correspond structurally with the mammalian paraventricular (PVN) and supraoptic (SON) nuclei. Although the distribution of AVT, identified by immunohistochemistry of adjacent serial sections within the hypothalamus, was similar to the distribution of AVT mRNA, the possibility that some positive staining was due to mesotocin neurons was not excluded. Water deprivation for 2 and 4 days resulted in both an increase in levels of AVT mRNA per neuron and the number of AVT mRNA-containing cells. Additionally, water deprivation resulted in a decrease in the amount of IR-AVT per neuron. The results indicate that osmotic stimulation increases AVT gene expression not only in individual neurons but also by activating subpopulation of neurons that are not observed in normally hydrated birds.

The effect of different temporal relationships of 5‐hydroxytryptophan (5‐HTP) and l‐dihydroxyphenylalanine (L‐DOPA) on reproductive and metabolic responses of migratory red‐headed bunting (Emberiza bruniceps)

Abstract To study the reproductive and metabolic effects of circadian serotonergic and Dopaminergic activities, 5‐hydroxytryptophan (5‐HTP, serotonin precursor) and L‐dihydroxyphenylalanine (L‐DOPA, dopamine precursor) were administered daily in different temporal relationships in Red‐headed bunting (Emberiza bruniceps) during the preparatory phase of annual gonadal cycle. L‐DOPA given daily after 12 hours of 5‐HTP injection (12‐hr relationship) induced full gonadal development and fat deposition. But, L‐DOPA given after 4 hours of 5‐HTP administration (4‐hr relationship) stimulated only fat deposition without any gonadal effect. On the other hand, these two drugs injected 8‐hour apart led to significant decrease in gonadal growth as well as fat deposition. The findings manifested that 12 hour relationship of these two neural oscillations may induce breeding conditions (summer), 8 hour relationship induced post reproductive regressive/photorefractory condition (late summer) and 4‐hour relationship induced...

Thyroidectomy and gonadal development in common myna, Acridotheres tristis

Abstract Thyroidectomy inhibited the gonadal development in the common myna, during quiescent, progressive, and reproductive phases. But, when performed during late reproductive and regressive phases it had no effect on the annual gonadal cycle.

Temporal Phase Relation of Circadian Neural Oscillations Alters RFamide-Related Peptide-3 and Testicular Function in the Mouse

In order to study the effect of the temporal synergism of neural oscillations on reproductive regulation and the response of RFamide-related peptide-3 (RFRP-3; a mammalian ortholog of avian gonadotropin-inhibitory hormone), expression of immunoreactive RFRP-3 in the neurons of the dorsomedial nucleus of the hypothalamus was monitored in sexually immature and mature laboratory mice (study I). In study II, the effects of serotonin and dopamine precursors (5-hydroxytryptophan and L-dihydroxyphenylalanine; injected daily, 8 or 12 h apart, for 13 days in 3-week-old mice) on testicular activity and immunoreactive RFRP-3 neurons were studied until 24 days after treatment. Results indicate high levels of expression of immunoreactive RFRP-3 in the sexually immature and 8-hour mice (simulating gonadal suppression), while a low level was noted in mature and 12-hour mice (simulating gonadal stimulation). These findings not only suggest the modulation of gonadal development in mice (during the course of puberty attainment) by changing the temporal phase relation of serotonergic and dopaminergic oscillations (as in some seasonally breeding species), but also demonstrate an inverse correlation of RFRP-3 neurons and gonadal activity in both control and experimental conditions.

Expression of hypothalamic arginine vasotocin gene in response to water deprivation and sex steroid administration in female Japanese quail

SUMMARY Arginine vasotocin (AVT) is a neurohypophyseal hormone involved in reproductive function and control of osmoregulation in birds. In view of the dual function of AVT, the present experiment was designed to observe the effect of water deprivation (WD) and sex steroid [estradiol benzoate (EB) and testosterone propionate (TP)] treatment independently, as well as simultaneously, on the profile/activity of the hypothalamic AVT system. WD resulted in a significant increase in plasma osmolality, sodium ion concentration and AVT concentration, but administration of sex steroids had no significant influence on these parameters. By contrast, the amount of hypothalamic AVT transcript (northern analysis) and the size of immunoreactive vasotocin (ir-AVT) neurons and hybridization signals (in the form of silver grains), representing AVT mRNA in corresponding neurons of paraventricular nuclei (PVN), increased significantly in all the treated groups compared with controls. Our findings indicate that although sex steroid administration has no effect on plasma osmolality and AVT concentration, unlike water deprivation, it may stimulate the profile/activity of AVT neurons of PVN, supporting the possibility of sex steroid receptors on these neurons. It is concluded that in quail, osmotic stress not only upregulates the expression of the AVT gene in existing neurons but also recruits many more neurons to increase the rate of AVT synthesis and secretion, while sex steroids appear to have a stimulatory effect only on the existing number of neurons and only at the level of transcription/translation and hence may influence/modulate hypothalamic AVT gene expression in response to osmotic stress. This study also suggests an interrelationship between reproduction and AVT system/function in birds.

Temporal synergism of neurotransmitter affecting drugs and seasonal reproductive responses of Indian Palm Squirrel,Funambulus pennanti

Daily injections of L-dihydroxyphenylalanine (L-DOPA, dopamine precursor) given 12 hour after 5-hydroxytryptophan (5-HTP, serotonin precursor) eliminated annual testicular regression in seasonally breeding sub-tropical Palm Squirrel that undergoes seasonal changes in responsiveness to day-length and humidity. Other temporal relations (L-DOPA given 0, 4, 8, 16 and 20 hours after 5-HTP administration) decreased/delayed the rate of regression and maintained the reproductive system at intermediate level. 12-hour-relation maintained full breeding condition (maximum gonad and accessory sex organs) unlike control, which exhibited complete atrophy of primary and accessory sex organs. It seems likely that these injections entrained the circadian serotonergic and dopaminergic oscillations and the interaction/phase relation of these two systems through their many circadian expressions (neural/hormonal) accounts for the seasonal interpretation of environmental factors. It is suggested that response in the present study is not a pharmacological effect of drugs but temporal synergism of neural acitivities affected the seasonality of reproduction as only 12-hour-relation of 5-HTP and L-DOPA maintained breeding activity during post reproductive phase of annual gonad cycle in Palm Squirrel.

2.45-GHz microwave irradiation adversely affects reproductive function in male mouse, Mus musculus by inducing oxidative and nitrosative stress

Abstract Electromagnetic radiations are reported to produce long-term and short-term biological effects, which are of great concern to human health due to increasing use of devices emitting EMR especially microwave (MW) radiation in our daily life. In view of the unavoidable use of MW emitting devices (microwaves oven, mobile phones, Wi-Fi, etc.) and their harmful effects on biological system, it was thought worthwhile to investigate the long-term effects of low-level MW irradiation on the reproductive function of male Swiss strain mice and its mechanism of action. Twelve-week-old mice were exposed to non-thermal low-level 2.45-GHz MW radiation (CW for 2 h/day for 30 days, power density = 0.029812 mW/cm2 and SAR = 0.018 W/Kg). Sperm count and sperm viability test were done as well as vital organs were processed to study different stress parameters. Plasma was used for testosterone and testis for 3β HSD assay. Immunohistochemistry of 3β HSD and nitric oxide synthase (i-NOS) was also performed in testis. We observed that MW irradiation induced a significant decrease in sperm count and sperm viability along with the decrease in seminiferous tubule diameter and degeneration of seminiferous tubules. Reduction in testicular 3β HSD activity and plasma testosterone levels was also noted in the exposed group of mice. Increased expression of testicular i-NOS was observed in the MW-irradiated group of mice. Further, these adverse reproductive effects suggest that chronic exposure to nonionizing MW radiation may lead to infertility via free radical species-mediated pathway.

Arginine vasotocin gene expression in hypothalamic neurons is up-regulated in chickens drinking hypertonic saline: an in situ hybridization study.

Osmotic stress stimulates the release of the avian hypothalamic neuropeptide arginine vasotocin (AVT) into the peripheral circulation. We conducted the present study to investigate the effects of salt-loading on AVT secretion and AVT gene expression in specific hypothalamic nuclei in chickens. White Leghorn chickens were provided food ad lib and either water or 2% NaCl to drink. Both plasma osmolality and plasma AVT levels were significantly increased in chickens that drank 2% NaCl for either two or four days compared to that in chickens that drank water. Results from in situ hybridization analysis demonstrated an increase in the number of neurons expressing AVT mRNA in the supraoptic (SON) and paraventricular nuclei (PVN) in chickens provided 2% NaCl to drink compared to chickens that were provided water to drink. The number of grains per neuron increased in the PVN, but not in the SON of osmotically stimulated birds. Thus, increased osmolality resulting from ingestion of hypertonic saline is an effective stimulus to increase hypothalamic AVT mRNA content in chickens.

Temporal synergism of neurotransmitters (serotonin and dopamine) affects testicular development in mice

The temporal phase relation of circadian oscillations is reported to regulate reproduction in many seasonally breeding avian and mammalian species, but its role in the reproductive regulation of continuous breeders is not yet known. Hence in the present study, six experimental groups of 3-week-old male Parkes strain mice, Mus musculus, were injected with 5-hydroxytryptophan (5-HTP, serotonin precursor) and L-dihydroxyphenylalanine (L-DOPA, dopamine precursor) at intervals of 0, 4, 8, 12, 16 or 20 hr (5mg/100g body weight per day for 13 days). Control mice received two daily injections of normal saline. When observed 24 days post-treatment, 8-hr mice exhibited low body weight and suppression of gonadal activity (spermatogenesis, sperm count/motility/viability and plasma testosterone concentration), while body weight and degree of gonadal development were higher in the 12-hr mice as compared to the controls. It is concluded that normal somatic and gonadal growth of pre-puberal mice may be suppressed with an 8-hr phase relation of circadian serotonergic and dopaminergic oscillations. On the other hand, a 12-hr phase relation accelerated the rate of gonadal maturation, while other relations led to more or less similar gonadal development as in the control mice. This study suggests the importance of circadian organization as a function of specific temporal phase relations of neural oscillations in the maturation of gonads. Although the exact mechanism still needs to be investigated, this seems to be mediated via effects on the neuroendocrine axis.

Apoptosis-mediated testicular alteration in Japanese quail (Coturnix coturnix japonica) in response to temporal phase relation of serotonergic and dopaminergic oscillations

ABSTRACT Reproductive performance of many avian species, including Japanese quail, is reported to be modulated by specific temporal phase relation of serotonergic and dopaminergic oscillations. Accordingly, it has been shown that the serotonin precursor 5-HTP and the dopamine precursor l-DOPA given 8 h apart induce gonadal suppression and given 12 h apart lead to gonadal stimulation, while other temporal relationships were found to be ineffective. In the present study, we investigated the effects of 8- and 12-h phase relation of neural oscillations on testicular responses including expression of GnRH-I, GnIH, pro-apoptotic proteins (p53 and Bax), inactive and active executioner caspase-3, and the uncleaved DNA repair enzyme PARP-1. Testicular volume and mass decreased significantly in 8-h quail and increased in 12-h quail compared with controls. Expression of ir-GnIH, p53, Bax and active-caspase-3 increased and that of GnRH-I, pro-caspase-3 and uncleaved PARP-1 decreased in 8-h quail compared with controls. A TUNEL assay also confirmed testicular regression in these quail. Testes of 12-h quail exhibited significantly increased expression of GnRH-I, pro-caspase-3 and uncleaved PARP-1 compared with the control group. Our findings suggest that differential response of avian testes to 8- and 12-h phase relation of serotonergic and dopaminergic neural oscillations may be attributed to autocrine/paracrine action of GnIH expression, which is upregulated in regressed testes, leading to apoptotic changes, and downregulated in developed testes, causing apoptotic inhibition. It is concluded that specific phase relation of neural oscillations may modulate the local testicular GnRH-GnIH system and alter the apoptotic mechanism in quail testes. Moreover, these findings highlight the physiological effects of time-dependent drug delivery, including the specific time intervals between two drugs. Summary: Testicular regression in quail in response to 8-h phase relation of 5-HTP and l-DOPA is GnIH induced, p53 dependent and Bax mediated, and this pathway reverses in hypertrophied 12-h quail testes.