Nitric oxide mediated kisspeptin regulation of steroidogenesis and gametogenesis in the catfish, Clarias batrachus.

Nitric oxide (NO) is a gaseous molecule that regulates various reproductive functions. It is a well-recognized regulator of GnRH-FSH/LH-sex steroid secretion in vertebrates including fish. Kisspeptin is a recently discovered neuropeptide which also regulates GnRH secretion. Nitrergic and kisspeptin neurons are reported in close physical contact in the mammalian brain suggesting their interactive role in the release of GnRH. The existence of kisspeptin and NOS is also demonstrated in vertebrate gonads, but information on their reciprocal relation in gonads, if any, is obscure. Therefore, attempts were made to evaluate the functional reciprocal relation between nitric oxide and kisspeptin in the catfish gonads, if any, by administering the nitric oxide synthase (NOS) inhibitor, L-NAME {N(G)-nitro-L-arginine methyl ester}, which reduces NO production, and kisspeptin agonist (KP-10) and assessing their impacts on the expressions of kisspeptin1, different NOS isoforms, NO and steroid production in the gonadal tissue. The results revealed that L-NAME suppressed the expression of kiss1 in gonads of the catfish establishing the role of NO in kisspeptin expression. However, KP-10 increased the expression of all the isoforms of NOSs (iNOS, eNOS, nNOS) and concurrently NO and steroids in the ovary and testis. In vitro studies also indicate that kisspeptin stimulates the production of NO and estradiol and testosterone levels in the gonadal explants and medium. Thus, in vivo results clearly suggest a reciprocal interaction between kisspeptin and NO to regulate the gonadal activity of the catfish. The in vitro findings further substantiate our contention regarding the interactive role of kisspeptin and NO in gonadal steroidogenesis.

Role of Neurokinin B in gametogenesis and steroidogenesis of freshwater catfish, Clarias batrachus.

Neurokinin B (NKB), a recently discovered neuropeptide, plays a crucial role in regulating the kiss-GnRH neurons in vertebrate's brain. NKB is also characterized in gonadal tissues; however, its role in gonads is poorly understood. Therefore, in the present study, the effects of NKB on gonadal steroidogenesis and gametogenesis through in vivo and in vitro approaches using NKB antagonist MRK-08 were evaluated. The results suggest that the NKB antagonist decreases the development of advanced ovarian follicles and germ cells in the testis. In addition, MRK-08 further reduces the production of 17ß-estradiol in the ovary and testosterone in the testis under both in vivo and in vitro conditions in a dose-dependent manner. Furthermore, the in vitro MRK-08 treatment of gonadal explants attenuated the expression of steroidogenic marker proteins, i.e., StAR, 3ß-HSD, and 17ß-HSD dose-dependently. Moreover, the MAP kinase proteins, pERK1/2 & ERK1/2 and pAkt & Akt were also downregulated by MRK-08. Thus, the study suggests that NKB downregulates steroidogenesis by modulating the expressions of steroidogenic marker proteins involving ERK1/2 & pERK1/2 and Akt/pAkt signalling pathways. NKB also appears to regulate gametogenesis by regulating gonadal steroidogenesis in the catfish.

Asprosin promotes steroidogenesis and spermatogenesis with improved glucose metabolism in adult mice testis.

Asprosin is an orexigenic adipokine that regulates appetite and glucose homeostasis in mammals. To date, only fragmentary findings are reported regarding its role in testicular activities. In the current investigation, immunolocalization and direct action of asprosin in adult mice testis was evaluated. Immunohistochemical and immunoblot studies were performed to analyse the testicular expression of asprosin. Intratesticular treatment of asprosin (0.1 µg and 1.0 µg per testis) was given to evaluate its direct action on testicular functions. Sertoli and Leydig cells were found to be immuno-positive for asprosin. Intratesticular administration of asprosin resulted into a significant increase in glucose and lactate levels along with enhanced expression of asprosin receptor OLFR734, insulin receptor (IR), glucose transporter 8 (GLUT 8), lactate dehydrogenase (LDH) activity and monocorboxylate transporters (MCT2 and 4). In addition, asprosin administration increased the testicular expression of cell proliferation (proliferating cell nuclear antigen: PCNA), cell survival (B cell lymphoma 2: Bcl2) and decreased germ cell apoptosis (Cysteine aspartic acid protease 3: Caspase 3) leading to increased sperm counts. Further, asprosin treatment resulted into increased level of total cholesterol, testosterone and steroidogenic markers (steroidogenic acute regulatory protein: StAR; 3beta-hydroxysteroid dehydrogenases: 3ß HSD and 17beta-hydroxysteroid dehydrogenases: 17ß HSD). Asprosin treatment promotes testicular glucose uptake and lactate synthesis to provide energy for steroidogenesis and spermatogenesis. The significant correlation between the asprosin-induced increased IR expression and increased testosterone, glucose and lactate levels suggests its role in increased survival and proliferation but decrease in germ cell apoptosis. This study proposed asprosin's role as an autocrine/paracrine regulator of testicular functions in adult mice.

Sub-chronic restraint stress exposure in adult rats: An insight into possible inhibitory mechanism on testicular function in relation to germ cell dynamics.

Psychological stress is now widely recognized as one of the major risk factors for male fertility. Its impact on the dynamics of testicular germ cells, however, has yet to be fully investigated. Therefore, we used the rat restraint stress (RS) model as a psychological stressor to assess the impact of psychological stress on testicular germ cell dynamics. Adult male SD rats were exposed to sub-chronic RS for 1.5 and 3 h per day for 30 days. The quality of cauda epididymis spermatozoa was adversely affected by RS exposure, and the frequency of spermatozoa with tail abnormalities was higher than that of spermatozoa with head abnormalities. RS exposure adversely affected testicular daily sperm production by disturbing the meiotic and post meiotic germ cell kinetics in the testis. The histomorphology of the testis was altered by loosening and vacuolization in the seminiferous epithelium, germ cell exfoliation and the presence of giant cells. Seminiferous tubules of stage I-VI and VII-VIII were severely affected in rats exposed to RS for 3 h. By interfering with steroidogenic enzymes, RS exposure disrupts testosterone biosynthesis. The testicular oxidative balance was also disturbed by RS exposure, which disrupted the levels/activities of lipid peroxidation, Nrf-2, superoxide dismutase and catalase. There was also an increase in caspase-3 activity and a decrease in the Bax-Bcl2 ratio. In conclusion, our findings suggest that psychological stressors like RS impair testicular functions in rats by disrupting germ cell dynamics, downregulating testicular androgenesis and increasing oxidative stress and apoptosis.

Photo-thermal regulation of neuropeptide Y (NPY) expression in ovarian follicles and ovarian activity of the catfish, Clarias batrachus.

Authors have recently reported a gradual increase in neuropeptide Y expression in the ovarian follicles of Clarias batrachus with the progression of oogenesis, coinciding with increasing photoperiod and temperature. This indicates the involvement of photoperiod and temperature in controlling NPY expression. Therefore, a study was designed to investigate the role of photoperiod and temperature in regulation of NPY expression in ovarian follicles. The catfish were exposed to different photo-thermal regimes during the late-quiescence and late-recrudescence phases for one month, and the expression of NPY was analyzed along with other ovarian activities. Though the exposure of catfish to long photoperiod induced a marginal increase (1.5 fold) in NPY expression in follicular cells, the high temperature stimulated its expression more effectively (6-10 fold), irrespective of photoperiodic exposures. Exposure to long photoperiod and high temperature together induced NPY expression maximally in granulosa and thecal cells of fully grown oocytes, but exposure to low temperature decreased its expression significantly. The oogenic and steroidogenic activities were also promoted simultaneously after the exposure to high temperature and long photoperiod alone or in combination. However, the low temperature exposure suppressed the ovarian activities leading to atresia of advanced follicles. Thus it is suggested that photoperiod and temperature both affect NPY expression and ovarian recrudescence in fish but the influences of temperature seem to be more prominent.

Application of phytoremediation technology in decontamination of a fish culture pond fed with coal mine effluent using three aquatic macrophytes.

In the present study, three aquatic macrophytes, Eichhornia crassipes, Salvinia molesta, and Pistia stratiotes were used to assess their relative efficacies in decontamination of a fish culture pond, regularly fed with coal mine effluent (CME). The level of metals like Fe, Mn, Ni, Zn, Cu, Pb, Cr, and Cd were much higher in CME-fed pond water than their recommended limits in drinking water set by the Bureau of Indian standards and in effluents by the Environmental Protection Agency. The levels of metal were lowered substantially in CME-fed pond water after exposure of the above plants to such water, however, metal levels in the plants increased tremendously. The increased metal levels in plants severely damaged their physiological and biochemical processes. The contents of chlorophyll a, b and carotenoid were reduced by 63.2, 64.2, and 46.3%, respectively, in E. crassipes, 41, 57.4, and 57.8% in S. molesta, and 42, 62, and 61% in P. stratiotes. The accumulating metals also generated oxidative stress in plants, as evident from the increased superoxide dismutase and catalase activities and enhanced malondialdehyde content. The E. crassipes was the most potent in absorbing the metals from the CME-fed pond water, followed by S. molesta and P. stratiotes.

Coal mine effluent-led bioaccumulation of heavy metals and histopathological changes in some tissues of the catfish Clarias batrachus.

Coal mining generates huge quantity of toxic effluent which consistently pollutes the neighboring wetlands where the local inhabitants regularly cultivate edible fishes. In the present study the concentration of heavy metals Fe, Zn, Cu, Mn, Ni, Cd, Pb and Cr were analyzed in the water and various tissues of edible catfish Clarias batrachus reared in a pond receiving effluents from Rajrappa coal mine, Jharkhand, India. The metal concentrations in the pond water were dramatically higher (Fe 350%, Zn 423%, Cu 12%, Mn 7029%, Ni 713%, Cd 1700%, Pb 4333% and Cr 588%) than the safe limit of Environmental Pollution Agency (2003) as well as the control tap water. Excessive amounts of metals in effluent caused their substantial transfer to the different tissues of the catfish reared in such ponds. Results showed that accumulation of metals in fish tissues were in the following order: liver > kidney > air breathing organ (ABO) > gills > skin > brain > muscles. Among the various tissues the highest accumulation of most of the metals was recorded in the liver (2.05-271.28 mg/kg dry weight) and lowest in the muscles (1.39-30.27 mg/kg dry weight), while the concentration of metals in other tissues ranged in between. The accumulation of heavy metals in tissues appears to cause remarkable histopathological alterations in skin, gills, ABO, liver and kidney that might be leading to deleterious effect on fish physiology and consequently impact the consumers of such fishes.

Seasonal ovarian immunolocalization of neuropeptide Y and its role in steriodogenesis in Asian catfish, Clarias batrachus.

The present study was undertaken to examine the cellular localization and potential steroidogenic role of neuropeptide Y (NPY) in the ovary of the freshwater catfish, Clarias batrachus. NPY-immunoreaction was observed in the follicular cells (granulosa and thecal cells) in the growing ovarian follicles, and the intensity of staining increased steadily from the initiation of follicular development until follicles were fully grown. Thereafter as follicles matured the stain intensity decreased. Positive correlations were found between NPY expression and the ovarian levels of 17ß-estradiol, testosterone, and activities of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and 17ß-hydroxysteroid dehydrogenase (17ß-HSD) in the ovary. In vitro NPY treatment stimulated the production of the two steroids and the activities of two enzymes. This is the first report of NPY immunoreactivity at the cellular level in the fish ovary and implicates this orexigenic peptide in the modulation of ovarian steroidogenesis.

Nitric oxide (NO) stimulates steroidogenesis and folliculogenesis in fish.

The present study was undertaken to understand the physiological significance of the existence of nitric oxide synthase (NOS)/nitric oxide (NO) system in fish ovary. For this, two doses of NO donor, sodium nitroprusside (SNP, 25 µg and 50 µg) and NOS inhibitor, N-nitro-l-arginine methyl ester (l-NAME, 50 µg and 100 µg)/100 g body weight were administered during the two reproductive phases of reproductive cycle of the Clarias batrachus During the late-quiescence phase, high dose of l-NAME decreased the NO, testosterone, 17ß-estradiol, vitellogenin contents in serum and ovary and activities of 5-ene-3ß-hydroxysteroid dehydrogenases (3ß-HSD) and 17ß-hydroxysteroid dehydrogenases (17ß-HSD) in ovary, whereas higher dose of SNP increased these parameters. l-NAME also reduced oocytes-I but increased perinucleolar oocytes in the ovary, whereas SNP treatment increased the number of advanced oocytes (oocytes-I and II) than the perinucleolar oocytes when compared with control ovary. During the mid-recrudescence phase, both doses of SNP increased NO, testosterone, 17ß-estradiol and vitellogenin in serum and ovary; however, l-NAME treatment lowered their levels. The activities of ovarian 3ß-HSD and 17ß-HSD were also stimulated by SNP, but l-NAME suppressed their activities compared to the control. The SNP-treated ovaries were dominated by oocyte-II and III stages, whereas l-NAME-treated ovary revealed more perinucleolar oocytes and oocytes-I and practically no advanced oocytes. Expression of endothelial NOS (eNOS), inducible NOS (iNOS) and neuronal NOS (nNOS) was augmented by the SNP and declined by l-NAME treatments as compared to the control. This study, thus, provides distinct evidence of NO-stimulated steroidogenesis, vitellogenesis and folliculogenesis in fish.

Pro-steroidogenic and pro-spermatogenic actions of nitric oxide (NO) on the catfish, Clarias batrachus: An in vivo study.

In an earlier study we have demonstrated reproductive-stage dependent, cell specific existence of inducible nitric oxide synthase (iNOS) and neuronal nitric oxide synthase (nNOS)/NO system in testis of the catfish, Clarias batrachus. The present study is an extension to examine the role of NO in steroidogenesis and spermatogenesis through in vivo administration of a NO donor, sodium nitroprusside (SNP) and a NOS inhibitor, N-nitro-l-arginine methyl ester (l-NAME) during the quiescence and recrudescence phase of the reproductive cycle of the catfish. Effects of these chemicals were assessed on the gonadosomatic index (GSI), levels of circulating & testicular testosterone, NO, activities of 3ß-hydroxysteroid dehydrogenase (3ß-HSD), 17ß-hydroxysteroid dehydrogenase (17ß-HSD) in testis, expression of different NOS isoforms and testicular morphology in relation to spermatogenesis. SNP treatment increased the GSI, testicular and circulating testosterone & NO, activities of testicular 3ß-HSD & 17ß-HSD, and expression of NOS isoforms. It also increased the area and perimeters of interstitium and seminiferous tubules in the testis. It accelerated the spermatogenesis, as was evident from the large number of spermatids/spermatozoa in seminiferous tubules and very few spermatogonial cells/primary spermatocytes in comparison to the control testis. On the contrary, l-NAME significantly suppressed GSI, testosterone & NO levels in serum and testis, and activities of testicular 3ß-HSD & 17ß-HSD. It also suppressed the expression of NOSs in testis. Though l-NAME did not alter the spermatogonial mitotic proliferation with the advancement of testicular recrudescence, it halted the progression of spermatogenesis (meiotic division and spermatozoa formation) as was clear from the increase in spermatogonial cells and very few advanced germ cells in the seminiferous tubules in l-NAME treated testis, compared to the control testis. The above noted effects were highly pronounced in the recrudescing catfish. Their effects were very marginal and at a particular dose levels of SNP and l-NAME in the quiescent testis. This study distinctly provides evidence of pro-steroidogenic and pro-spermatogenic role of NO. This study also demonstrates the existence of eNOS in fish testis for the first time. The positive feedback control of expression of all isoform of NOS in testis by NO is also noteworthy.

Immunolocalization of nitric oxide synthase (NOS) isoforms in ovarian follicles of the catfish, Clarias batrachus and its relation with ovarian activity.

Nitric oxide, a gaseous molecule, is produced during the conversion of arginine to citrulline by the action of NOS isoforms (eNOS, iNOS or nNOS). Role of NO in regulation of mammalian reproduction is well established; however, practically no report is available on fishes. Hence, in the present study, expression of all three isoforms of NOS was worked out in the ovary of Clarias batrachus immunohistochemically during different phases of its reproductive cycle and its relation with ovarian activities. No immunoreactivity of eNOS was observed in the ovary of C. batrachus during the late-quiescence and early-recrudescence phases. While during the recrudescence phase (April and May) it expressed intensely in thecal and granulosa cells of the oocyte-II and III, but immune-intensity decreased in the late-recrudescence and spawning phases (June and July). Similar pattern of immunoprecipitation was also observed in case of iNOS. However, the immunoreactivity pattern of nNOS was quite varied, it expressed moderately only in the nucleus and cytoplasm of perinuclear and oocyte-I stages during late-quiescence phase. While during the early recrudescence phase, the expression of nNOS disappeared completely from the nucleus and cytoplasm, rather it expressed intensely in the thecal and granulosa cells, which declined in the late-recrudescence and spawning phases. Moderate immunoreactivity of iNOS could also be localized in the zona radiata of ovulated oocyte. The intense NOS immunoreactivity in the thecal and granulosa cells coincided with increased levels of ovarian NO and 17ß-estradiol content. They exhibited statistically significant positive correlation amongst themselves, suggesting the involvement of ovarian NOS/NO system in oogenesis and steroidogenesis in the catfish.

Temperature dependent action of growth hormone on somatic growth and testicular activities of the catfish, Clarias batrachus.

Effects of growth hormone on somatic growth and testicular activities were studied during late quiescence and early recrudescence phases of the reproductive cycle of the catfish, Clarias batrachus. The administration of exogenous growth hormone (GH) during the late quiescence phase (December-January; ambient water temperature-15.2±1°C) did not influence the somatic growth as well as the testicular activity, as no change in body weight, testis weight, plasma level of insulin-like growth factor I (IGF-I) and testicular morphology was detected following GH treatment, though the plasma testosterone was marginally increased. While during the early recrudescence phase (March-April; ambient water temperature-28.1±2°C), GH treatment promoted the production of insulin like growth factor-I and testicular steroidogenic activity in a dose dependent manner, as was evident from the significant increase in the circulating levels of testosterone and estradiol-17ß. GH treatment also increased body weight, testicular weight and gonadosomatic index, suggesting its involvement in testicular development. The GH treatment promoted spermatogonial proliferation and accelerated the spermatogenic process in the present catfish. These results, thus, suggest that GH influences the somatic growth and testicular activities depending on the temperature of the rearing water; warmer temperature and longer photoperiod promote testicular steroidogenic and spermatogenic activities in fish. This study has immense practical use in fisheries science.

Derivation and characterization of a ES-like cell line from indian catfish Heteropneustes fossilis blastulas.

A cell line designated as HFB-ES was established from blastula stage embryos of H. fossilis (Singhi). The embryonic cells were harvested and maintained in Leibovitz's medium supplemented with 15% fetal bovine serum. The cell line had been subcultured for more than 90 passages in a period of 24 months. HFB-ES cells were able to grow at temperatures between 25 and 35°C with an optimum temperature of 28°C. The growth rate of HFB-ES was proportional to FBS concentration, with optimum growth seen at 15% FBS concentration. The originality of the cell line was confirmed by sequencing of cytochrome oxidase c subunit I (COI), cytochrome b gene, and microsatellite DNA profile. Results of chromosome complements of HFB showed normal karyo-morphology with 56 (2n) diploid number of chromosomes after 40 passages which indicated that the developed cell line is chromosomally stable. The pluripotency of HFB was demonstrated by alkaline phosphatase activity and Oct-4 gene expression. Expression of GFP reporter gene was successful in HFB-ES. These results indicated that HFB-ES could be utilized for future gene expression studies.

Season-dependent effect of thermopulse on gonadal recrudescence in the female catfish, Clarias batrachus.

In the present study, the female Clarias batrachus, held under long photoperiod (13L:11D), were exposed to high water temperature either constantly (24 h) and/or in form of thermopulse of 6 h and 12 h durations, separately, at different times of the day/night cycle for six weeks during the early post-spawning and late post-spawning phases of its reproductive cycle. The effects of high water temperature (30 +/- 1 degrees C) on gonadosomatic index (GSI), plasma levels of testosterone (T) and oestradiol-17beta (E2) were observed. During the late post-spawning phase, thermopulse of 12 h duration given in the morning hour increased all the studied parameters most effectively as compared to that given at evening hour of the day/night cycle or even in comparison to the fish exposed to constant high temperature. Thermopulse of 6 h duration given in the morning or noon also raised these parameters compared to the controls, but the magnitudes of stimulation were moderate. However, exposures of the catfish to such photothermal regimes during the early post-spawning phase completely failed to bring any change in the studied parameters. These findings, thus, clearly indicate that treatment with high temperature under long photoperiod may stimulate gonadal activity in C. batrachus, provided given at appropriate season of the year. A diurnal basis of response to high temperature and the existence of a rigid gonado-refractory phase (perhaps just after the spawning) are also evident in the reproductive cycle of C. batrachus.

Malathion exposure induces the endocrine disruption and growth retardation in the catfish, Clarias batrachus (Linn.).

Many hormones are known for their role in the regulation of metabolic activities and somatic growth in fishes. The present study deals with the effects of malathion (an organophosphorous pesticide) on the levels of metabolic hormones that are responsible for promotion of somatic and ovarian growth of the freshwater catfish, Clarias batrachus. Malathion treatment for thirty days drastically reduced the food intake and body weight of fish. These fish also exhibited a great avoidance to food. Exposure of catfish to malathion reduced the levels of thyroxine (T(4)), triiodothyronine (T(3)), growth hormone (GH), insulin like growth factor-I (IGF-I), testosterone (T) and estradiol-17ß (E(2)) in a dose dependent manner during all the studied reproductive phases, in general, except that malathion increased the level of GH during the quiescence phase. Significant reduction in muscle and hepatic protein content also occurred in the malathion-treated fish. Malathion exposure induced lipolysis too in the liver and muscle. The results thus support that malathion treatment disrupts the endocrine functions and the olfactory sensation responsible for food intake and gustatory feeding behavior, which ultimately leads to retardation of fish growth.

Analyses of the health status, risk assessment and recovery response of the nutritionally important catfish Clarias batrachus reared in coal mine effluent-fed pond water: a biochemical, haematological and histopathological investigation.

The present field study evaluates the health status of the catfish Clarias batrachus reared in coal mine effluent (CME)-fed pond water at Rajrappa mining complex using biochemical, haematological and histopathological parameters. Simultaneously, risk assessment along with recovery response of the CME intoxicated fish following their treatment with CME-free freshwater was also studied. The CME-fed pond water fish revealed significant decrease in biomolecules concentrations and considerable increase in activities of several enzymes along with metallothionein level as compared to control. The impaired regulation of metabolic function was also revealed by blood parameters showing significant decrease in haemoglobin content (8.78 ± 0.344 g/100 mL) and red blood cells count (1.77 ± 0.12 × 106 mm3) while substantial elevation in white blood cells (187.13 ± 9.78 × 103 mm3). The histopathological study also confirmed the changes including hypertrophy of club cells of skin, swelling of secondary lamella of gills, extensive fibrosis in liver and glomerular shrinkage with increased Bowman's space in kidney. Potential health risk assessments based on estimated daily intake and target hazard quotient indicated health risks associated with the consumption of such fishes. The CME-contaminated fish when transferred to CME-free freshwater exhibited decreased metal content accompanied by eventual recovery response as evident by retrieval in biochemical and haematological parameters. Withdrawal study also revealed restoration in the activity of different marker enzymes in fish tissues including blood as well as recovery in their cellular architecture. The results of the present study validate the depuration process as an effective practice for detoxification of fish contaminated with effluent.

Coal mine effluent-induced metal bioaccumulation, biochemical, oxidative stress, metallothionein, and histopathological alterations in vital tissues of the catfish, Clarias batrachus.

In the present study, a multi-biomarker approach was used to assess the toxicity of the coal mine effluent (CME) generated at the Rajrappa coal mine on the catfish Clarias batrachus. A core of biomarkers indicative of nutritional value, oxidative stress, and histopathology was selected to illustrate the toxic effects of CME-containing different heavy metals and other toxicants. The results of metal bioaccumulation in CME-exposed fish tissues revealed the highest metal concentration in liver (1.34-297.68 mg/kg) while lowest in muscles (1.47-23.26 mg/kg) as compared to other tissues and so was the metallothionein level. The high value of bioaccumulation observed in liver, kidney, and gills reflects their affinity for metals. In addition, the values of metal pollution index (MPI) of different fish tissues further affirmed that liver followed by kidney and gills are at greater risk than brain, skin, and muscles. Significant alterations in the activity of certain enzymes (aspartate amino transferase, alanine amino transferase, alkaline phosphatase) as well as oxidative stress markers (superoxide dismutase, catalase and lipid peroxidation) were detected in the tissues of CME-exposed fish. The tissue-specific metal accumulation and increased metallothionein levels may be associated with the biochemical and physiological activity of an organ and its constitutive antioxidant defenses. The histopathological changes in the various tissues of the CME-exposed fish justify the high metal accumulation and biochemical alterations. Overall results indicate that the Rajrappa coal mine effluent is very toxic having adverse health impact on the fish and might also affect the human health when consumed.

Seasonal expression and distribution of kisspeptin1 (kiss1) in the ovary and testis of freshwater catfish, Clarias batrachus: A putative role in steroidogenesis.

The central role of kisspeptin (kiss) in mammalian reproduction is well established; however, its intra-gonadal role is poorly addressed. Moreover, studies investigating intra-gonadal role of kiss in fish reproduction are scanty, contradictory and inconclusive. The expression of kiss1 mRNA has been detected in the fish brain, and functionally attributed to the regulation of reproduction, feeding and behavior. The kiss1 mRNA has also been demonstrated in tissues other than the brain in some studies, but its cellular distribution and role at the tissue level have not been adequately addressed in fish. Therefore, an attempt was made in the present study to localize kiss1 in gonadal cells of the freshwater catfish, Clarias batrachus. This study reports the presence of kiss1 in the theca cells and granulosa cells of the ovarian oocytes and interstitial cells in the testis of the catfish. The role of kiss1 in the ovary and testis of the catfish was also investigated using kiss1 receptor (kiss1r) antagonist (p234). The p234 treatment decreased the production of 17ß-estradiol in ovary and testosterone in the testis by lowering the activities of 3ß-hydroxysteroid dehydrogenase and 17ß-hydroxysteroid dehydrogenase under both, in vivo as well as in vitro conditions. The p234 treatment also arrested the progression of oogenesis, as evident from the low number of advancing/advanced oocytes in the treated ovary in comparison to the control ovary. It also reduced the area and perimeter of the seminiferous tubules in the treated catfish testis. Thus, our findings suggest that kiss is involved in the regulation of gonadal steroidogenesis, independent of known endocrine/ autocrine/ paracine regulators, and thereby it accelerates gametogenic processes in the freshwater catfish.

Gametogenic and steroidogenic action of kisspeptin-10 in the Asian catfish, Clarias batrachus: Putative underlying mechanistic cascade.

Unlike mammals, two kisspeptins genes encoding, kiss1 and kiss2 are detected in fishes with highly varied and contradictory difference in their reproductive activities. The present study was undertaken to examine the direct action of kisspeptin-10 and its role in gonadal activities in the gonadally quiescent Asian catfish using native mammalian kisspeptin decapeptide (KP-10) involving in vivo and in vitro approaches. The in vivo KP-10 treatment caused precocious onset of gametogenesis and its rapid progression, as was evident from the appearance of advanced stages of ovarian follicles in ovary, and advanced germ cells (spermatocytes/ spermatids) in the testis of the treated Clarias batrachus in comparison to the control gonads. It also elevated the steroid levels in gonads of the catfish in vivo and in vitro conditions. Simultaneously, it increased the expressions of key steroidogenic enzymes like 3ß-HSD, 17ß-HSD, and StAR protein, responsible for transfer of cholesterol from outer to inner membrane of the mitochondria of steroidogenic cells. Concurrently, it augmented the activities of 3ß-HSD and 17ß-HSD in the ovarian explants. The expressions of MAPK component (pERK1/2 and ERK1/2) were also up-regulated by KP-10 in gonadal explants. Thus, the data suggest that kisspeptin-10 stimulates gametogenesis by enhancing gonadal steroid production. The study also describes the putative mechanistic cascade of steroidogenic actions of kisspeptin-10 in the catfish so much so in teleost fish. The study also suggests that, kisspeptin may act locally to regulate gonadal activities in an autocrine/paracine manner, independent of known extra-gonadal factors in the catfish.