Chronic stress induced duration dependent alterations in immune system and their reversibility in rats.

The objective was to find out whether severity of stress effects on immunity increases with duration of exposure and recovery depends on duration of exposure. Adult male rats (n = 30) were subjected to restraint (1 h) followed by forced swimming exercise (15 min) after a gap of 4 h daily for 2, 4 and 8 weeks and allowed to recover for 6 weeks after each exposure period. Exposure of rats to stress resulted in duration dependent significant decreases in leukocyte count, phagocytic indices of neutrophils, number of bone marrow stem cells and serum levels of IL-12 and increases in apoptotic index of peripheral blood mononuclear cells and serum levels of IL-10. The alterations in counts of neutrophils, total immunoglobulin content, phagocytic index, apoptotic index of peripheral blood mononuclear cells and serum levels of IL-10 returned to control levels in recovery group rats of 2 and 4 weeks exposure but not in that of 8 weeks exposure. However, alterations in number and apoptotic index of bone marrow stem cells returned to control levels in 2, 4 and 8 weeks stress recovery groups. The results for the first time reveal that increase in duration of exposure results in more severe damage in immune system and that shorter the exposure period, faster the recovery. In addition, in vitro study for the first time showed that corticosterone causes apoptosis of peripheral blood mononuclear cells and bone marrow stem cells in dose dependent manner. Hence death of leukocytes and their stem cells is the major cause of stress induced immune dysfunction.

Chronic stress effects and their reversibility on the Fallopian tubes and uterus in rats.

The durational effects of chronic stress on the Fallopian tubes and uterus were studied by exposing rats to stressors in the form of restraint (1h) and forced swimming (15min) daily for 4, 8 or 12 weeks. One group of stressed rats from each time period was then maintained without exposure to stressors for a further 4 weeks to assess their ability to recover from stress. All time periods of stress exposure resulted in decreased weight of the body and Fallopian tubes; however, the relative weight of the uterus and serum concentrations of oestradiol and insulin increased significantly. The antioxidant potential was decreased with increased malondialdehyde concentrations in the Fallopian tubes following all durations of exposure and after 4 and 8 weeks of stress exposure in the uterus. Interestingly, rats stressed for 12 weeks showed an increase in serum testosterone concentration and antioxidant enzyme activities with a decrease in malondialdehyde concentration in the uterus. The antioxidant enzyme activities and malondialdehyde concentration in the Fallopian tubes of all recovery group rats were similar to stressed rats. However, in the uterus these parameters were similar to controls in recovery group rats after 4 weeks or 8 weeks of exposure, but after 12 weeks of stress exposure these parameters did not return to control levels following the recovery period. These results reveal, for the first time, that chronic stress elicits an irreversible decrease in antioxidant defence in the Fallopian tubes irrespective of exposure duration, whereas the uterus develops reversible oxidative stress under short-term exposure but increased antioxidant potential with endometrial proliferation following long-term exposure.

The protective effect of the Vacha rhizome extract on chronic stress-induced immunodeficiency in rat.

CONTEXT: Chronic stress is an inevitable factor in the modern day society which affects cell mediated as well as humoral immunity. There is a need to prevent stress effects with traditionally used herbs. OBJECTIVE: The present study was undertaken to investigate the immunoprotective effect of Vacha (Acorus calamus L. Acoraceae) rhizome under stressful condition. MATERIALS AND METHODS: Soxlet extraction of Vacha rhizome was performed with increasing polarity of solvents, i.e., petroleum ether to ethanol. The extract was concentrated by distilling off the solvent in flash evaporator and dried in desiccators. The benzene extract was found to have anti-stress property in our earlier studies and hence it was used in the present experiment. Extract was administered every day for 4 weeks orally to adult female rats prior to exposure to stress, restraint (1 h) and forced swimming exercise (15 min). RESULTS: Vacha rhizome extract significantly prevented the stress induced reduction in total and differential leukocytes count, immunoglobulin content, bone marrow cellularity and viability, lymphocytes counts in lymphoid organs, islands of white pulp of spleen (ED50 = 10 mg, p < 0.001) and a significant increase in circulating immune complexes and apoptotic index of lymphoid organs (ED50 = 10 mg, p < 0.001) compared to controls. DISCUSSION AND CONCLUSION: The present study clearly indicates that Vacha extract not only prevents stress-induced suppression of immunity and structural involution of lymphoid organs, but also boosts immunity in normal rats. Therefore, it is suggested that Vacha extract administration maintains normal immunity despite the body experiencing stress.

Long-term chronic stress exposure induces PCO phenotype in rat.

Thus far the effects of chronic stress on the ovary were studied for shorter durations. However, responses of the ovary may vary with durations of exposure to stress. Hence, we investigated the responses of the ovary following exposure to different durations of chronic stress. Exposure of rats to restraint (1 h) and after a gap of 4 h to forced swimming (15 min) daily for 4 or 8 weeks resulted in significant decreases in the activities of the ovarian antioxidant enzymes, 3ß-hydroxysteroid dehydrogenase and percentage of healthy granulosa cells with concomitant increases in the number of atretic follicles, the percentage of apoptotic granulosa cells and ovarian malondialdehyde concentration. However, the response of the ovary to similar stress regime for 12 weeks was paradoxical as there were increases in the activities of ovarian antioxidant enzymes and 3ß-hydroxysteroid dehydrogenase, the number of healthy antral follicles, and decreases in ovarian malondialdehyde concentration and percentage of apoptotic granulosa cells. These changes were accompanied by hyperglycaemia and an increase in the serum levels of insulin, testosterone and oestradiol. In addition the cystic follicles were found in the ovaries of these rats. However, the number of oestrous cycles and active corpora lutea showed significant decrease in all the durations of stress exposure. The results demonstrate a differential response of ovary to short- and long-term exposure to chronic stress.

Progesterone prevents corticosterone mediated inhibition of estrous behaviour in rats.

Stress induced by application of electric foot shocks (300 microA/shock, five shocks per episode, 4 episodes at 1800, 1830, 1900 and 1930 hrs on the proestrus day) to rats at the time of pre-ovulatory progesterone secretion, abolished lordosis and resulted in maximum rejection co-efficient, whereas treatment with a CRF receptor antagonist (alpha-helical CRF9-41) or metapirone, an inhibitor of corticosterone synthesis, prior to application of the electric foot shocks, resulted in normal lordosis and a significant reduction in rejection coefficient. Further, administration of a single dose of corticosterone (40 microg) at 1800 hrs of proestrus caused inhibition of lordosis and resulted in maximum rejection co-efficient. On the other hand, corticosterone + progesterone treatment at 1800 hrs of proestrus resulted in normal lordosis and a significant reduction in rejection coefficient. The facts that stress induced inhibition of lordosis is prevented by CRF receptor antagonist or metapirone and that corticosterone inhibits lordosis indicate that stress induced inhibition of lordosis is mediated by corticosterone. Further, normal display of lordosis by rats treated with corticosterone + progesterone in contrast to its absence in corticosterone alone treated rats suggests that impaired progesterone secretion due to action of corticosterone leads to inhibition of lordosis.

Progesterone restores stress induced inhibition of estrous behavior in rat.

Exposure to a stressor (mild electrical shocks to foot, five times per episode, at 1800, 1830, 1900 and 1930 hrs of proestrus) coinciding with period of pre-ovulatory progesterone secretion in rats abolished estrous behavior as shown by the absence of lordosis response and a significant increase in rejection quotient compared to controls. These rats did not show spermatozoa in the vaginal smear next day morning in contrast to their presence in controls. On the other hand, rats treated with progesterone (a single injection, 500 microg in 0.1 ml olive oil at 1800 hr of proestrus) prior to exposure to stressor showed normal estrous behavior, as shown by significantly lower rejection quotient than rats exposed to stress alone, lordosis quotient similar to controls and presence of spermatozoa in the vaginal smear next day. The results, albeit indirectly, to the best of our knowledge, first time indicate that stress induced impaired steroidogenesis leads to suppression of estrous behavior.

Light microscopical and ultrastructural studies on the vas deferens of the lizard Mabuya carinata.

Adult male lizards (Mabuya carinata) were studied during breeding and non breeding seasons to determine the regional and seasonal differences if any in the vas deferens and to compare ultrastructural features of luminal epithelial cells with those of endotherms. The vas deferens of the lizard is a convoluted tube extending from the epididymis to the hemipenis passing over the kidney. Based on morphometric data of luminal diameter and epithelial cell height three distinct regions viz; proximal, middle and distal regions were identified in the vas deferens. The epithelium is surrounded by a thin layer of lamina propria, many layers of circular smooth muscle fibers and an outer layer of visceral pleuro peritoneum. Based on cell and nuclear morphology and ultrastructure, five different cell types viz; principal cell, basal cell, mitochondria rich cell, halo cell and narrow cell were identified in the epithelium during both breeding and non breeding season. Principal cells and basal cells were more abundant in both seasons. The types of luminal epithelial cells of vas deferens of M. carinata and their ultrastructural features are similar to those of mammals. Further, vas deferens of M. carinata differs from mammals in having only circular smooth muscles in contrast to circular and longitudinal muscles of mammalian vas deferens. To the best of our knowledge this is the first report describing cell types of vas deferens, their ultrastructure and ultrastructural seasonal variations in reptiles.

Stress inhibits seasonal and FSH-induced ovarian recrudescence in the lizard,Mabuya carinata.

Stressors (handling, chasing, and noise) applied randomly five times per day for one month to lizards during the recrudescence phase of the ovarian cycle caused a significant reduction in mean number of oocytes and primordial follicles when compared to those of controls. Further, vitellogenic follicles were absent in the ovary of lizards subjected to stressors. Administration of bovine FSH during post-breeding regression phase of the ovarian cycle induced ovarian recrudescence as shown by significant increases in the mean number of oogonia, oocytes, and primordial follicles compared to controls, as well as vitellogenic growth of follicles. However, lizards treated with FSH and exposed to stressors did not exhibit ovarian recrudescence. Furthermore, FSH administration during the post-breeding regression phase caused a significant increase in serum levels of estradiol compared to controls, which was accompanied by significant increases in the relative weight of the liver and oviduct, as well as vitellogenic growth of follicles. Despite administration of FSH to lizards subjected to stressors, there was neither any increase in serum levels of estradiol and weight of the liver nor vitellogenic growth of follicles. The results indicate that repeated application of stressors inhibits vitellogenic growth of follicles by suppression of steroidogenic activity in M. carinata. This is the first report revealing that the ovary does not respond to gonadotrophin treatment under stressful conditions in reptiles.

Influence of corticosterone on FSH-induced ovarian recrudescence in the lizard Mabuya carinata.

Administration of bovine FSH (10 IU/lizard/alternate day for 30 days) in the postbreeding quiescent phase of the ovarian cycle caused a significant increase in the mean number of oogonia and oocytes, the relative weight of the oviduct, and the liver and serum estradiol levels compared to those of controls. In addition, the FSH-treated lizards showed a vitellogenic growth of follicles and development through to preovulatory follicles. However, the administration of corticosterone simultaneously with FSH (10 IU FSH + 40 microgram corticosterone/lizard/alternate day for 30 days) did not result in these changes and the ovaries resembled those of controls. The results indicate the absence of ovarian refractoriness to gonadotropic stimulation during the quiescent phase of the reproductive cycle and inhibition of FSH-induced ovarian recrudescence by corticosterone. It is suggested that corticosterone treatment reduces FSH-induced steroidogenic activity of the ovary, leads to impairment in vitellogenin secretion by the liver, and results as well in the failure of vitellogenic follicular growth in Mabuya carinata.

LHRH induced changes in the interrenal gland of the frog Rana cyanophlyctis (Schn.).

Administration (i.m.) of synthetic mammalian LHRH (40 ng/frog, alternate day for 30 days) to adult male intact frogs R. cyanophlyctis resulted in activation of the interrenal cells as shown by increases in nuclear diameters of the interrenal cells, activity (histochemical demonstration) of delta 5-3 beta-hydroxysteroid dehydrogenase and glucose-6-phosphate dehydrogenase and concomitant decrease in sudanophilic lipid droplets in the interrenal gland when compared to those of controls. These changes were not observed in the interrenal gland of LHRH treated hypophysectomized frogs. The results indicate that LHRH stimulates secretory activity of the interrenal cells and action may not be directly on the adrenal gland.

Histophysiological studies on sex steroid induced changes in the interrenals of the male frog Rana cyanophlyctis (Schn.).

Administration (i.m.) of either estradiol-17 beta (0.5 microgram/frog) or progesterone (10 micrograms/frog) on alternate days for 15 days to adult male frogs induced the following changes in the interrenal cells: i) nuclear and cellular hypertrophy, ii) increase in the activity of delta 5-3 beta-hydroxy-steroid dehydrogenase and iii) decrease in sudanophilic lipid droplets. Treatment with testosterone propionate (100 micrograms/frog) dit not elicit marked changes in the above parameters. These results demonstrate that some sex steroids modify the interrenal gland steroidogenic activity in R. cyanophlyctis.

Effects of metopirone on the pars distalis, the interrenal gland and the testis of the frog, Rana cyanophlyctis (Schn.).

Administration (i.m.) of metopirone (2.5 mg/frog/day for 10 days) to adult male frog, R. cyanophlyctis, resulted in marked degranulation and hypertrophy of B3 cells and a moderate degranulation of B2 cells of the pars distalis. Concomitantly, there was hypertrophy of the interrenal cells and regression of the Leydig cells, however spermatogenesis was not affected. The results suggest a functional correlation between pituitary B cells and the interrenals in R. cyanophlyctis.