Kappa-opioid receptor-mediated modulation of innate immune response by dynorphin in teleost Channa punctatus
Introduction
Innate immune response is crucial for the survival of lower vertebrates as the adaptive immune response takes time to be operational. The phagocytic cells, importantly macrophages and neutrophils, are the major components of innate immune system and form the first line of host defense mechanism. They remove pathogens by phagocytosis or releasing cytotoxic substances. There are numerous studies in mammals that evidence the involvement of endogenous opioid peptides in regulation of phagocyte functions such as phagocytosis [6], [16], [25], [27], [36] and production of nitric oxide [5], [13], [21], [34], [38], superoxide [5], [28], [33], [35], [38], tumor necrosis factor-α [17] and interleukins [3], [4]. Also, the immunomodulatory role of β-endorphin and methionine-enkephalin through specific opioid receptor has been studied in a few fish [31], [32], [39]. Nonetheless, the immunoregulatory property of dynorphin, an endogenous opioid peptide, has not been explored so far in ectotherms including fish.
It is worth mentioning that cDNA for prodynorphin cloned from fish shows a higher degree of similarity with that of other vertebrates [12], [14], [1]. In mammals, the prodynorphin gene product contains a dynorphin-A(32), dynorphin-A(1–17), dynorphin-A(1–8), dynorphin-B(29), and α- and β-neoendorphins. The prodynorphins in non-mammalian vertebrates contain an additional opioid peptide, which is methionine-enkephalin in the amphibian Bufo marinus [10], Leucine-enkephalin in the sacropterigian fish Neoceratodus forsteri [12] and Ile-enkephalin in actinopterigian fish [1]. Moreover, the zebrafish prodynorphin contains the consensus sequences for four opioid peptides: an Ile-enkephalin, the α- and β-neoendorphins, dynorphin-A(1–17) and dynorphin-B(29). Out of these, dynorphin-A(1–17) is the most abundant form, and it differs with human dynorphin-A(1–17) at positions 5 and 13 [14]. Although sequencing of dynorphin-A(1–17) is not done so far in Channa punctatus, its sequence available in other fish like eel (Anguilla rostrata) and lungfishes (Australian lungfish N. forsteri and African lungfish Protopterus annectens) differs only at two to three positions from that of human [1], [12]. Nonetheless, N-terminal region is found to be highly conserved and responsible for the receptor binding activity [12], [22]. Hence, the present study was aimed to investigate the role of dynorphin-A(1–17) in regulation of non-specific immune responses, e.g. phagocytosis, superoxide and nitrite production in a teleost C. punctatus. Further, an attempt has also been made to find out the involvement of specific opioid receptor through which dynorphin exerts its effect on phagocyte functions.
Section snippets
Animals
Adult male fish C. punctatus weighing 80–100 g were procured from neighboring state of Delhi (latitude 28.38°N, longitude 77.2°E) and acclimated to the laboratory conditions for fortnight under 12 L:12 D light regimens at 25 ± 2 °C. They were fed on alternate day with minced beef liver. The guidelines of the committee for the purpose of control and supervision of experiments on animals (CPCSEA), Ministry of Statistics and Programme Implementation, Government of India were followed for use and
Concentration-related effect of dynorphin-A(1–17)
The data analyzed by ANOVA showed concentration-related marked variation (P < 0.001) in dynorphin-A(1–17) effect on phagocytosis, superoxide production and nitrite release by splenic phagocytes of C. punctatus. In general, dynorphin-A(1–17) had stimulatory effect on phagocytosis and superoxide production, while inhibitory effect on nitrite release. In case of phagocytosis, a significant (Newman–Keuls’ multiple range test, P < 0.05) increase was noticed at 10−13 M concentration, the lowest
Discussion
In the present study, an endogenous opioid peptide dynorphin-A(1–17) shows the immunomodulatory role in teleost C. punctatus. Dynorphin-A(1–17), the most prevalent form in fish, increased the phagocytosis of yeast cells Saccharomyces cerevisiae by splenic phagocytes. The present finding is parallel to that observed with β-endorphin [32] and methionine-enkephalin [31] on phagocytosis in our earlier studies in C. punctatus. The stimulatory effect of endogenous opioid β-endorphin on phagocytic
Acknowledgments
The first author is grateful to Council of Scientific and Industrial Research, India for fellowship. Authors are thankful to Indian Council of Agricultural Research (ICAR), PUSA, New Delhi, India for financial support. Authors are indebted to National Institute on Drug Abuse (NIDA), Bethesda, USA, for the kind gift of dynorphin-A(1–17) and selective opioid receptor antagonists.
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