u-50488 and Hypertrophy

To observe the effect of κ-opioid receptor (κ-OR) stimulation on Angiotensin II (Ang II)-induced cardiomyocyte hypertrophy in vitro cultured myocardial cells from neonatal rats and on calcineurin (CaN) signal pathways.. Cultured myocardial cells of neonatal rats were divided into control group, CSA (1 µmol/L) group, Ang II (1 µmol/L) group, Ang II (1 µmol/L) + U50488H (1 µmol/L) group, Ang II (1 µmol/L) + CSA (1 µmol/L) group, Ang II (1 µmol/L) + Rp-cAMPS (1 µmol/L) group, Ang II (1 µmol/L) + CSA (1 µmol/L) + U50488H (1 µmol/L) group and Ang II (1 µmol/L) + PTX5 mg/L + U50488H (1 µmol/L) group. The hypertrophic myocytes were induced by Ang II 1µmol/L before κ-OR agonist U50488H 1 µmol/L was administered. The antihypertrophic effect of κ-OR stimulation was observed in the presence of ciclosporine A (CsA) 1 µmol/L, cAMP triethyl-ammonium salt (Rp-cAMPS) 1 µmol/L, and pertussistoxin ( PTX) 5 mg/L. The total protein content was assayed by the method of Lowry. The [Ca²⁺]i was measured by confocal microscope using Fluo-3/AM as flouresecent indicator. The relative expression of CaN was determined by Western blot.. (1) The total protein content of Ang II group was significantly higher than that in control group (P<0.01), which could be equally reduced by cotreatment with U50488H, CSA and Rp-cAMPS (P<0.01). Total protein content of the Ang II + PTX + U50488H group and the Ang II group was similar. (2) The [Ca²⁺]i was significantly higher in Ang II group of neonatal rat cardiomyocytes than that in control group (P<0.01), which could be reduced by cotreatment with U50488H, CSA and Rp-cAMPS (P<0.01). [Ca(2+)]i was similar between the Ang II + PTX + U50488H group and the Ang II group. (3) The expression of CaN was significantly higher in Ang II group than that in control group (P<0.01), which could be significantly reduced by cotreatment with U50488H, CSA and Rp-cAMPS (P<0.01). CaN was similar between the Ang II + PTX + U50488H group and the Ang II group.. κ-opioid receptor activation could attenuate Ang II induced cardiomyocytes hypertrophy via reducing [Ca²⁺]i and downreglating CaN.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Angiotensin II; Aniline Compounds; Animals; Animals, Newborn; Calcineurin; Cells, Cultured; Hypertrophy; Myocardium; Myocytes, Cardiac; Rats; Receptors, Opioid, kappa; Signal Transduction; Xanthenes

We aimed to further define the pathway mediating the inhibitory effects of kappa-opioid receptor stimulation on Ca(2+) transients and hypertrophic responses to beta(1)-adrenoceptor stimulation. We determined the effects of trans-(+/-)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]-benzeneacetamid methanesulfonate salt (U50,488H), a selective kappa-opioid receptor agonist, on the enhancement of spontaneous Ca(2+) transients and the induction of hypertrophy by isoprenaline, a beta-adrenoceptor agonist, in cultured neonatal ventricular myocytes. The results were compared with those found with KN93, a selective Ca(2+)/calmodulin-dependent kinase (CaMKII) inhibitor, propranolol, a beta-adrenoceptor antagonist, and verapamil, a L-type Ca(2+) channel antagonist. Hypertrophy of cardiomyocytes was characterized by increases in (i) total protein content; (ii) cell size; and (iii) [(3)H]leucine incorporation. 10 micromol/l isoprenaline increased all three parameters. We also determined the expression of nuclear CaMKIIdelta in response to U50,488H in the presence or absence of isoprenaline. To determine whether the effects of U50,488H were receptor-mediated, its effects were also measured following blockade of the kappa-opioid receptor with nor-binaltorphimine. kappa-Opioid receptor stimulation suppressed the stimulatory effect of isoprenaline on Ca(2+) transients and cardiac hypertrophy, as did KN93, propranalol and verapamil. U50,488H also suppressed the expression of nuclear CaMKIIdelta(B) in the presence, but not in the absence of isoprenaline. These results suggest that the inhibitory effect of kappa-opioid receptor stimulation on beta(1)-adrenoceptor stimulation may also involve CaMKIIdelta.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Adrenergic beta-Agonists; Adrenergic beta-Antagonists; Animals; Animals, Newborn; Benzylamines; Calcium Channel Blockers; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Nucleus; Cell Size; Cells, Cultured; Hypertrophy; Isoproterenol; Myocytes, Cardiac; Naltrexone; Narcotic Antagonists; Propranolol; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, beta-1; Receptors, Opioid, kappa; Sulfonamides; Verapamil