Abstract
Prodynorphin mRNA and dynorphin B expression have been previously shown to be greatly increased in cardiac myocytes of BIO 14.6 cardiomyopathic hamsters. Here we report that exogenous dynorphin B induced a dose-dependent increase in prodynorphin mRNA levels and stimulated prodynorphin gene transcription in normal hamster myocytes. Similar responses were elicited by the synthetic selective kappa opioid receptor agonist U-50,488H. These effects were counteracted by the kappa opioid receptor antagonist Mr-1452 and were not observed in the presence of chelerythrine or calphostin C, two specific protein kinase C (PKC) inhibitors. Treatment of cardiomyopathic cells with Mr-1452 significantly decreased both prodynorphin mRNA levels and prodynorphin gene transcription. In control myocytes, dynorphin B induced the translocation of PKC-alpha to the nucleus and increased nuclear PKC activity without affecting the expression of PKC-delta, -epsilon, or -zeta. Acute release of either U-50,488H or dyn B over single normal or cardiomyopathic cells transiently increased the cytosolic Ca2+ concentration. A sustained treatment with each opioid agonist increased the cytosolic Ca2+ level for a more prolonged period in cardiomyopathic than in control myocytes and led to a depletion of Ca2+ from the sarcoplasmic reticulum in both groups of cells. The possibility that prodynorphin gene expression may affect the function of the cardiomyopathic cell through an autocrine mechanism is discussed.
Highlights
Prodynorphin mRNA and dynorphin B expression have been previously shown to be greatly increased in cardiac myocytes of BIO 14.6 cardiomyopathic hamsters
Using the same methodology outlined in our companion studies [50, 51], we have examined whether Dynorphin B (dyn B) or the synthetic selective opioid receptor agonist U-50,488H may affect the levels of prodynorphin mRNA in hamster ventricular myocytes
Since we observed only a slight further increase in prodynorphin mRNA levels following the exposure of cardiomyopathic myocytes to dyn B or to U-50,488H, we cannot exclude that the amount of dyn B being released by cardiomyopathic cells might have approached the maximal effect of the opioid ligand on the expression of the prodynorphin gene
Summary
Prodynorphin mRNA and dynorphin B expression have been previously shown to be greatly increased in cardiac myocytes of BIO 14.6 cardiomyopathic hamsters. The observation that opioid receptor stimulation is coupled to protein kinase C (PKC) [8] and that PKC is involved in the regulation of prodynorphin gene expression [4] raises the possibility that the gene itself may be regulated in an autocrine fashion by one of its peptide products In this regard, it may be conceivable that dyn B would affect prodynorphin gene expression in pathological conditions characterized by an increase in the synthesis and release of this opioid peptide from the myocardial cell. We aimed at investigating whether the stimulation of opioid receptors by dyn B or by U-50,488H, a synthetic selective opioid receptor agonist [9], may affect prodynorphin mRNA expression or the rate of transcription of the prodynorphin gene in cardiac myocytes isolated either from normal or from cardiomyopathic hamsters. The possible consequences of opioid receptor agonism were further investigated in normal and cardiomyopathic cells by examining the effects produced by a short- or long-term exposure to U-50,488H or dyn B on the cytosolic Ca2ϩ level and on the releasable sarcoplasmic reticular Ca2ϩ pool
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