Ovarian nesfatin-1 in Hemidactylus flaviviridis: Reproductive phase-dependent expression, role and hormonal regulation.

Nesfatin-1 has recently emerged as a modulator of ovarian functions in mammals. Studies in non-mammalian vertebrates, though limited and majorly restricted to fishes, have evidenced a role of this peptide in the regulation of ovarian steroidogenesis and oocyte maturation. Interestingly, nesfatin-1 remains completely unexplored in reptiles. Therefore, the present study aimed to identify nesfatin-1 and elucidate its role and regulation in the ovary of Hemidactylus flaviviridis. Ovarian expression of nucb2/nesfatin-1 was highest during late recrudescence and breeding while it was lowest during regression. Follicular stage-dependent expression analysis showed significantly high expression of nucb2/nesfatin-1 in previtellogenic follicles. Further, in vitro treatment of recrudescent wall lizard ovaries with nesfatin-1 resulted in increased expression of anti-apoptotic gene, bcl-2, along with a concomitant decline in the pro-apoptotic gene, caspase-3. In addition, proliferation/differentiation markers like scf, c-kit, pcna, and bmp-15 were stimulated in ovaries incubated with the peptide. Ovarian steroidogenesis was also positively influenced by nesfatin-1 as treatment with the peptide resulted in heightened star expression as well as increased estradiol and progesterone production. Also, all concentrations of nesfatin-1 stimulated glucose uptake and metabolism in wall lizard ovary. Our observations provide the first evidence of ovarian functions of nesfatin-1 in a reptile. Further, ovarian nucb2/nesfatin-1 was differentially regulated by gonadotropin and sex steroids wherein its expression was stimulated by dihydrotestosterone (DHT) and 17ß-estradiol (E2) but inhibited by follicle-stimulating hormone (FSH). In summary, this is the first report of the presence, reproductive stage-dependent expression, role, and regulation of ovarian nucb2/nesfatin-1 in H. flaviviridis.

Genotoxic Risks to Male Reproductive Health from Radiofrequency Radiation.

During modern era, mobile phones, televisions, microwaves, radio, and wireless devices, etc., have become an integral part of our daily lifestyle. All these technologies employ radiofrequency (RF) waves and everyone is exposed to them, since they are widespread in the environment. The increasing risk of male infertility is a growing concern to the human population. Excessive and long-term exposure to non-ionizing radiation may cause genetic health effects on the male reproductive system which could be a primitive factor to induce cancer risk. With respect to the concerned aspect, many possible RFR induced genotoxic studies have been reported; however, reports are very contradictory and showed the possible effect on humans and animals. Thus, the present review is focusing on the genomic impact of the radiofrequency electromagnetic field (RF-EMF) underlying the male infertility issue. In this review, both in vitro and in vivo studies have been incorporated explaining the role of RFR on the male reproductive system. It includes RFR induced-DNA damage, micronuclei formation, chromosomal aberrations, SCE generation, etc. In addition, attention has also been paid to the ROS generation after radiofrequency radiation exposure showing a rise in oxidative stress, base adduct formation, sperm head DNA damage, or cross-linking problems between DNA & protein.

Significance of Complement Regulatory Protein Tetraspanins in the Male Reproductive System and Fertilization.

Fertilization is a very sophisticated and unique process involving several key steps resulting in a zygote's formation. Recent research has indicated that some immune system-related cell surface molecules (CD molecules from the tetraspanin superfamily) may have a role in fertilization. Extracellular vesicles are undeniably involved in a variety of cellular functions, including reproduction. Tetraspanin proteins identified in extracellular vesicles are now used mostly as markers; mounting evidence indicates that they also participate in cell targeting, cargo selection, and extracellular vesicle formation. Their significance and potential in mammalian reproduction are currently being studied extensively. Despite the fact that the current data did not establish any theory, the crucial function of tetraspanins in the fertilization process was not ruled out, and the specific role of tetraspanins is still unknown. In this review, we bring insight into the existing knowledge regarding the expression of tetraspanins in spermatozoa and seminal fluid and their role in gamete binding and fusion.

The role of non-ionizing electromagnetic radiation on female fertility: A review.

With increasing technological developments, exposure to non-ionizing radiations has become unavoidable as people cannot escape from electromagnetic field sources, such as Wi-Fi, electric wires, microwave oven, radio, telecommunication, bluetooth devices, etc. These radiations can be associated with increased health problems of the users. This review aims to determine the effects of non-ionizing electromagnetic radiations on female fertility. To date, several in vitro and in vivo studies unveiled that exposure to non-ionizing radiations brings about harmful effects on oocytes, ovarian follicles, endometrial tissue, estrous cycle, reproductive endocrine hormones, developing embryo, and fetal development in animal models. Non-ionizing radiation also upsurges the free radical load in the uterus and ovary, which leads to inhibition of cell growth and DNA disruptions. In conclusion, non-ionizing electromagnetic radiations can cause alterations in both germ cells as well as in their nourishing environment and also affect other female reproductive parameters that might lead to infertility.

Seasonality, sex-specificity and transcriptional regulation of hepatic leptin system in spotted snakehead Channa punctata.

The present study deals with sex-specific reproductive phase-dependent variation and sex steroids-induced transcriptional regulation of hepatic lep and lepr in nutritionally valuable spotted snakehead, Channa punctata. The data on seasonality reveals sex-specific variation in pattern of lep transcription where a high level was recorded during resting and postspawning quiescent phases in female while during resting and spawning phases in male. Unlike lep, lepr exhibited similar expression pattern along the reproductive phases in both the sexes. As compared to female, a three-fold higher expression of lep was detected in male during reproductively active phase only. However, no sexual dimorphism was evidenced in lepr either during active or quiescent phase. To explore the implication of sex steroids in regulation of leptin system, we correlated levels of plasma testosterone (T) and 17ß-estradiol (E2) with leptin system in males as well as females. Further, criss-cross in vivo and in vitro experiments with dihydrotestosterone (DHT) and E2 were conducted in male and female spotted snakehead. The leptin system was downregulated after DHT administration in both the sexes. However, with E2, a marked decrease was evidenced in male only. The sex-wise variable response of leptin system to sex steroids was validated by in vitro experiments wherein liver fragments from male and female fish were incubated individually with both the sex steroids. In conclusion, sex steroids modulate hepatic leptin system differentially depending on sex and reproductive state of spotted snakehead.

Radiofrequency radiation: A possible threat to male fertility.

Radiofrequency exposure from man-made sources has increased drastically with the era of advanced technology. People could not escape from such RF radiations as they have become the essential part of our routine life such as Wi-Fi, microwave ovens, TV, mobile phones, etc. Although non-ionizing radiations are less damaging than ionizing radiations but its long term exposure effect cannot be avoided. For fertility to be affected, either there is an alteration in germ cell, or its nourishing environment, and RF affects both the parameters subsequently, leading to infertility. This review with the help of in vitro and in vivo studies shows that RF could change the morphology and physiology of germ cells with affected spermatogenesis, motility and reduced concentration of male gametes. RF also results in genetic and hormonal changes. In addition, the contribution of oxidative stress and protein kinase complex after RFR exposure is also summarized which could also be the possible mechanism for reduction in sperm parameters. Further, some preventative measures are described which could help in reverting the radiofrequency effects on germ cells.

Reproductive phase-dependent variation, sexually dimorphic expression and sex steroids-mediated transcriptional regulation of lep and lepr in lymphoid organs of Channa punctata.

The reproductive phase-dependent and sex-related differential expression of leptin (lep) and its receptor (lepr) in primary and secondary lymphoid organs of a highly nutritive economically important Channa punctata preempts the involvement of sex steroids in modulating intra-immuno-leptin system. This hypothesis was strengthened when plasma testosterone (T) and estradiol (E2) levels in male and female fish of reproductively active spawning and quiescent phases were correlated with lep and lepr expression in their immune organs. Splenic lep and lepr showed a negative correlation with T in both male and female, while with E2 there was a positive correlation in male and negative in female C. punctata. In head kidney, a contrasting correlation was observed as compared to spleen. To validate the implication of sex steroids in regulating leptin system in immune organs, in vivo and in vitro experiments were performed with DHT and E2. Upon administration, lep and lepr expression in tissues of either sex was downregulated. In addition, in vitro results with either of the sex steroids exemplified their direct involvement. Overall, this study, for the first time, reports correlation between sex steroids and transcript expression of leptin system in immune organs of a seasonally breeding vertebrate.

Temporal expression and gonadotropic regulation of aromatase and estrogen receptors in the ovary of wall lizard, Hemidactylus flaviviridis: Correlation with plasma estradiol and ovarian follicular development.

The current study in Indian wall lizard Hemidactylus flaviviridis for the first time demonstrates the reproductive phase-dependent expression pattern of aromatase (cyp19) and estrogen receptor subtypes (er-α and er-ß) as well as their gonadotropic regulation in the ovary of a squamate. The expression of cyp19 remained low during regressed phase, increased markedly in recrudescent and declined sharply in breeding phase. Further, temporal profile of plasma estradiol 17-ß (E2) was found to be relatively parallel to the expression pattern of ovarian cyp19. The expression pattern of estrogen receptors in the ovary showed subtype-specific variation along the reproductive cycle. Expression of ovarian er-α remained high from regressed to late recrudescence, while er-ß expression that was low during regression dramatically increased with the initiation of follicular growth in early recrudescence and remained high until late recrudescence. Nonetheless, expression of both the receptors declined during breeding phase when ovary contained vitellogenic follicle. Regarding gonadotropic regulation, short-term treatment with Follicle stimulating hormone (3 injections of FSH) increased the ovarian expression of cyp19, er-α and er-ß while prolongation of treatment (7 or 11 injections) resulted in a marked decrease in expression of these genes concomitant to formation of vitellogenic follicle. However, a marked increase in plasma E2 was recorded after 7 injections of FSH. The direct role of gonadotropin in regulation of cyp19 and estrogen receptors was established by an in vitro study where FSH upregulated the expression of these genes in all stages of ovarian follicles (early growing, previtellogenic and early vitellogenic) of wall lizards.

Seasonality of reproduction in male spotted murrel Channa punctatus: correlation of environmental variables and plasma sex steroids with histological changes in testis.

The present study was undertaken to develop a comprehensive understanding of how environmental cues and sex steroids relate with cyclic changes in spermatogenesis in freshwater spotted snakehead Channa punctatus that is nutritious and economically important. The seasonal histological changes in testis and annual profile of gonadosomatic index (GSI) of C. punctatus delineated the testicular cycle into four phases: regressed (December-March), preparatory (April-June), spawning (July and August) and postspawning (September-November). Among environmental variables, correlation and regression analyses exhibited an important relationship between photoperiod and testicular weight while role of rainfall was seen confined to spawning. The seasonal profile of plasma sex steroids when correlated with cyclic changes in spermatogenesis in spotted snakehead, testosterone (T) seems to be involved in controlling the major events of spermatogenesis from renewal of stem cells to spawning of spermatozoa. Another important androgen prevalent in teleosts, 11-ketotestosterone (11-KT), was high during preparatory phase, suggesting that 11-KT in addition to T plays an important role in progression of spermatogenesis and spermiation in C. punctatus. However, 11-KT was not seen to be associated with milt production and release of spermatozoa during spawning. Plasma profile of estradiol-17ß (E2) during different reproductive phases revealed the involvement of E2 in repopulation of stem cells during postspawning phase and in maintaining quiescence of testis during regressed phase.

A Study of Differential Expression of Testicular Genes in Various Reproductive Phases of Hemidactylus flaviviridis (Wall Lizard) to Derive Their Association with Onset of Spermatogenesis and Its Relevance to Mammals.

Testis of Hemidactylus flaviviridis, commonly known as Indian wall lizard, displays a lack of cellular and metabolic activity in regressed phase of testis during non-breeding season of the year. Retracted Sertoli cells (Sc), fibroid myoid cells and pre-meiotic resting spermatogonia are observed in such testis. This situation is akin to certain forms of infertility in men where hormone supplementation fails to generate sperm despite the presence of Sc and germ cells (Gc) in testis. In testis of lizard, spermatogenesis is reinitiated upon increased level of hormones during appropriate season (phase of recrudescence). Study of genes associated with generation of sperm, from regressed adult testis in lizard, may provide valuable information for understanding certain forms of male idiopathic infertility. Subtractive hybridization using testicular RNA obtained from the regressed and active phases of lizard reproductive cycle led to identify eight partial mRNA sequences that showed sequence homology with mice genes. We further evaluated the gene expression prolife by real-time PCR in three different reproductive phases of H. flaviviridis: regressed (pre-meiotic), recrudescent (meiotic) and active (post meiotic), for comparison with the corresponding testicular phases found in testis of 5 days (pre-meiotic), 20 days (meiotic) and 60 days (post-meiotic) old mouse. This is the first report where genes associated with progression of spermatogenesis during active phase, which follows a regressed state of adult testis, were identified in lizard and found to be conserved in mouse. Six important genes, Hk1, Nme5, Akap4, Arih1, Rassf7 and Tubb4b were found to be strictly associated with active spermatogenesis in both mouse and lizard. Factors interfering with the expression of any of these genes may potentially abrogate the process of spermatogenesis leading to infertility. Such information may shed light on unknown causes of idiopathic male infertility.

Quercetin supplementation restores testicular function and augments germ cell survival in the estrogenized rats.

Quercetin, as a flavonoid, has been recognized to possess dual properties of an oxidant and antioxidant as well. The role of quercetin (QC), as an antioxidant in countering estradiol-3-benzoate (EB) induced adverse effects and germ cell apoptosis in adult rat testis was presently investigated. Adult rats received EB (0.075 mg/rat/5th day) alone or EB+QC (15 mg/kg bw/alternate day) simultaneously for 30 days. Revival of spermatogenesis following QC intervention was associated with a significant restoration in serum and intra-testicular levels of testosterone. Decline in lipid peroxidation and simultaneous improvement in the activities of superoxide dismutase, catalase and glutathione s-transferase were very much evident. Identically, total antioxidant capacity and glutathione demonstrated a marked improvement. QC augmented germ cell survival leading to a decrease in cell apoptosis. Expression of downstream apoptotic markers, caspase-3 and poly-ADP-ribose polymerase (PARP) presented a significant reduction. Down regulation with respect to upstream markers, caspase-8 and -9, Fas, FasL, Bax, and p53 was similarly observed. Taken together, the above findings indicate that with the dose presently used quercetin with its antioxidant and antiestrogenic properties restored testicular function leading to revival of spermatogenesis. It also augmented germ cell survival primarily mediated through downregulation in the expressions of upstream, downstream and other markers in the pathways of metazoan apoptosis.

Clomiphene citrate potentiates the adverse effects of estrogen on rat testis and down-regulates the expression of steroidogenic enzyme genes.

OBJECTIVE: To investigate the antiestrogenic effect of clomiphene citrate (CC) in male rats estrogenized with estradiol-3-benzoate (EB). DESIGN: Prospective experimental study. SETTING: Laboratory. ANIMALS: Adult male albino rats (Holtzman strain). INTERVENTION(S): CC was given alone or in combination with EB. MAIN OUTCOME MEASURE(S): Testicular function and steroidogenic enzyme gene expression were evaluated in control versus treated groups. RESULT(S): EB after 30 days of treatment induced a rise in TUNEL-positive germ cells adversely affecting spermatogenesis with complete absence of elongated spermatids or sperms. CC alone had only a moderate effect. In contrast, CC+EB synergistically inflicted more adverse effects as apoptotic germ cells per tubule rose further. Significant down-regulation in expression of testicular steroidogenic enzyme genes StAR, p450scc, 3ß-HSD, and p450c17 was observed. In the EB-alone group, aromatase gene expression in the testis was up-regulated but reversed in brain and liver tissues. CC alone had little modulatory effect on aromatase expression. On the other hand, CC+EB countered the EB-induced rise of aromatase expression in the testis. CONCLUSION(S): The above findings indicate that CC in the presence of estrogen synergistically potentiates more adverse effects in testis, inhibiting expression of upstream steroidogenic enzyme genes and leading to disruption of steroidogenesis.

Antileukemic Potential of Momordica charantia Seed Extracts on Human Myeloid Leukemic HL60 Cells.

Momordica charantia (bitter gourd) has been used in the traditional system of medicine for the treatment of various diseases. Anticancer activity of M. charantia extracts has been demonstrated by numerous in vitro and in vivo studies. In the present study, we investigated the differentiation inducing potential of fractionated M. charantia seed extracts in human myeloid HL60 cells. We found that the HL60 cells treated with the fractionated seed extracts differentiated into granulocytic lineage as characterized by NBT staining, CD11b expression, and specific esterase activity. The differentiation inducing principle was found to be heat-stable, and organic in nature. The differentiation was accompanied by a downregulation of c-myc transcript, indicating the involvement of c-myc pathway, at least in part, in differentiation. Taken together these results indicate that fractionated extracts of M. charantia seeds possess differentiation inducing activity and therefore can be evaluated for their potential use in differentiation therapy for leukemia in combination with other inducers of differentiation.

Dynorphin regulates the phagocytic activity of splenic phagocytes in wall lizards: involvement of a κ-opioid receptor-coupled adenylate-cyclase-cAMP-PKA pathway.

This in vitro study of the wall lizard Hemidactylus flaviviridis demonstrates the role of the opioid peptide dynorphin A((1-17)) [dyn A((1-17))] in the regulation of the phagocytic activity of splenic phagocytes. Dyn A((1-17)) in a concentration-dependent manner inhibited the phagocytic activity, and the maximum inhibition was recorded at a concentration of 10(-9) mol l(-1). To explore the receptor-mediated effect of dyn A((1-17)), cells were treated simultaneously with the non-selective opioid receptor blocker naltrexone and dyn A((1-17)). Naltrexone completely blocked the inhibitory effect of dyn A((1-17)) on phagocytosis. Moreover, the involvement of selective opioid receptors was investigated using selective opioid receptor antagonists. CTAP and naltrindole, selective µ- and δ-opioid receptor blockers, respectively, failed to block the inhibitory effect of dyn A((1-17)) on phagocytosis. However, the selective κ-opioid receptor blocker NorBNI completely antagonized the inhibitory effect of dyn A((1-17)). Regarding the κ-opioid receptor-coupled downstream signaling cascade, the adenylate cyclase (AC) inhibitor SQ 22536 and protein kinase A (PKA) inhibitor H-89 decreased the inhibitory effect of dyn A((1-17)) on phagocytosis. Furthermore, treatment with dyn A((1-17)) caused an increase in intracellular cAMP content in splenic phagocytes. Thus, it can be concluded that, in H. flaviviridis, dyn A((1-17)) negatively regulates the phagocytic activity of splenic phagocytes by acting through κ-opioid receptors that are coupled with the AC-cAMP-PKA signal transduction mechanism.

Immunomodulatory role of substance P in the wall lizard Hemidactylus flaviviridis: an in vitro study.

Present in vitro investigation for the first time in ectotherms demonstrated the immunomodulatory role of substance P in the wall lizard Hemidactylus flaviviridis. Substance P inhibited the percentage phagocytosis and phagocytic index of lizard splenic phagocytes. Inhibitory effect of substance P was completely blocked by NK-1 receptor antagonist spantide I, indicating the NK-1 receptor mediated action. Further, NK-1 receptor-coupled downstream signaling cascade involved in controlling phagocytosis was explored using inhibitors of adenylate cyclase (SQ 22536) and protein kinase A (H-89). Both the inhibitors, in a concentration-related manner decreased the suppressive effect of substance P on phagocytosis. In addition, substance P treatment caused an increase in intracellular cAMP level in splenic phagocytes. Taken together, it can be suggested that substance P via NK-1 receptor-coupled AC-cAMP-PKA pathway modulated the phagocytic activity of splenic phagocytes in wall lizards.

Neuropeptide Y, an orexigenic hormone, regulates phagocytic activity of lizard splenic phagocytes.

Present in vitro study in the wall lizard Hemidactylus flaviviridis, for the first time in ectothermic vertebrates, demonstrated the immunoregulatory role of neuropeptide Y (NPY) and its receptor-coupled downstream signaling cascade. NPY inhibited the percentage phagocytosis and phagocytic index of splenic phagocytes. The inhibitory effect of NPY on phagocytosis was completely antagonized by Y(2) and Y(5) receptor antagonists. This suggests that NPY mediated its effect on phagocytosis through Y(2) and Y(5) receptors. Further, NPY receptor-coupled downstream signaling cascade for NPY effect on phagocytosis was explored using the inhibitors of adenylate cyclase (SQ 22536) and protein kinase A (H-89). The SQ 22536/H-89 in a concentration-related manner decreased the inhibitory effect of NPY on phagocytosis. Further, an increase in intracellular cAMP level was observed in response to NPY. Taken together, it can be concluded that NPY via Y(2) and Y(5) receptor-coupled AC-cAMP-PKA pathway downregulated the phagocytic activity of lizard splenic phagocytes.

ß-Endorphin inhibits phagocytic activity of lizard splenic phagocytes through µ receptor-coupled adenylate cyclase-protein kinase A signaling pathway.

The receptor-coupled intracellular signaling mechanism of endogenous opioid peptide ß-endorphin (ß-end) is explored for the first time in ectothermic vertebrates using wall lizard as a model. ß-End inhibited the percentage phagocytosis and phagocytic index of lizard splenic phagocytes in a dose-dependent manner. The inhibitory effect of ß-end on phagocytosis was completely antagonized by non-selective opioid receptor antagonist naltrexone and also by selective µ-receptor antagonist CTAP. However, selective antagonists for other opioid receptors like NTI for δ-receptor and NorBNI for κ-receptor did not alter the effect of ß-end on phagocytosis. This suggests that ß-end mediated its inhibitory effect on phagocytic activity of splenic phagocytes exclusively through µ opioid receptors. The µ opioid receptor-coupled downstream signaling cascade was subsequently explored using inhibitors of adenylate cyclase (SQ 22536) and protein kinase A (H-89). Both SQ 22536 and H-89 abolished the inhibitory effect of ß-end on phagocytosis in a concentration-related manner. Implication of cAMP as second messenger was corroborated by cAMP assay where an increase in intracellular cAMP level was observed in response to ß-end treatment. It can be concluded that ß-end downregulated the phagocytic activity of lizard splenic phagocytes through µ opioid receptor-coupled adenylate cyclase-cAMP-protein kinase A pathway.

Role of estrogen in the regulation of spermatogenesis in the Indian wall lizard Hemidactylus flaviviridis.

The role of estrogen in the Indian wall lizard, Hemidactylus flaviviridis during PMSG induced spermatogenesis in the regression phase and during normal spermatogenesis in the active breeding phase was investigated. Blood hormone levels demonstrated a high testosterone to estrogen ratio in the breeding and vice verse during the regressed phase. PMSG treatment (30 IU in 100 µl saline/lizard/alternate day for 30 days) during the regressed phase stimulated spermatogenesis which was associated with a significant (p<0.001) rise in plasma testosterone levels. Complete spermatogenesis with sperms was resolved in many tubular sections. However, co-administration of PMSG plus estrogen in high doses (2 µg of estradiol benzoate/alternate day) for the same period not only curtailed germ cell proliferation significantly but also induced apoptosis in germ cells. There was no significant reduction in testicular weight but sperms were found completely absent in all the tubules. Decline in the plasma testosterone was more pronounced in high compared to low estrogen treated groups. Further, low estrogen administration had little effect either on raising the plasma levels of estrogen or subsequently on spermatogenesis which was identically observed in the breeding phase too. Estrogen intervention (2 µg) in the breeding phase also profoundly suppressed spermatogenesis leading to a severe depletion in germ cells. Simultaneously, there was a significant rise in germ cell apoptosis which was associated with an up-regulation of extrinsic (caspase 8, Fas, FasL) and intrinsic (caspase 9, Bax, Bcl2) markers in these cells. Taken together, the above data indicate that the estrogen plays a key role in regulating spermatogenesis in the wall lizard retarding it during testicular quiescence and eliminating germ cells through apoptosis during the active breeding phase.

Kappa-opioid receptor-mediated modulation of innate immune response by dynorphin in teleost Channa punctatus.

The immunomodulatory role of endogenous opioid peptides released during stress has been extensively studied in mammals, but least explored in lower vertebrates. The present in vitro study for the first time reports the specific opioid receptor-mediated immunomodulatory role of dynorphin-A((1-17)) in ectotherms. Dynorphin-A((1-17)) had pleiotropic effects on phagocyte functions, stimulatory on phagocytosis and superoxide production while inhibitory on the nitrite release. However, the effect of dynorphin-A((1-17)), whether stimulatory or inhibitory, markedly declined at high (10(-5)M) concentration. Dynorphin-A((1-17)) seems to mediate its action through opioid receptors since non-selective opioid receptor antagonist, naltrexone, completely blocked the effect of dynorphin-A((1-17)) on phagocytosis, superoxide production and nitrite release. Moreover, among specific opioid receptors antagonists, only selective kappa (kappa)-opioid receptor antagonist norbinaltorphimine was capable to antagonize the pleiotropic effects on phagocyte functions. The present study provides the direct evidence of immunomodulatory role of dynorphin-A((1-17)) via kappa-opioid receptor in freshwater teleost Channa punctatus.

Genomic and non-genomic effect of cortisol on phagocytosis in freshwater teleost Channa punctatus: an in vitro study.

The non-genomic action of glucocorticoid, besides classical genomic action, is recently implicated in regulation of phagocyte activities in mammals. With regard to the non-mammalian vertebrates, this study in the teleost, Channa punctatus, for the first time demonstrates the regulation of innate immunity by cortisol following non-genomic pathway. Cortisol suppressed the phagocytic activity of splenic phagocytes in a time- and concentration-dependent manner. Intriguingly, it impeded the phagocytosis within 15 min which is too short for conventional genomic action. The cortisol-induced rapid inhibition could not be altered by transcription and translation inhibitors, suggesting the involvement of non-genomic pathway. Since membrane impermeable BSA-cortisol mimicked the rapid inhibitory effect of cortisol at 15 min, we speculated that cortisol exerted its non-genomic effect on phagocytosis by acting at membrane site. These membrane-bound glucocorticoid receptors seem similar to cytosolic GR, as rapid inhibitory effect of cortisol was blocked by the cytoplasmic glucocorticoid receptor blocker RU-486. Using inhibitors for adenylate cyclase/protein kinase A (PKA) and estimating intracellular cAMP, adenylate cyclase-PKA pathway was seen involved in mediating the rapid non-genomic action of cortisol in phagocytes of C. punctatus. In contrast to the rapid effect, inhibitory effect of cortisol on phagocytosis after 1h was blocked by protein synthesis inhibitors, thus implicating genomic regulation. An overview of our data suggests that cortisol regulates phagocytosis in C. punctatus via genomic as well as non-genomic mechanisms. Further, the non-genomic action of cortisol is mediated via membrane-bound GR coupled to cAMP-PKA system.