Aiming to determine if sarcolenimal Ca^2+ transport system(s) in cardiac cells are functionally altered in diabetes, we investigated the responsiveness to extracellular Ca^2+ Ca^2+ _o ) of papillary ...muscles and ventricular myocytes isolated from rats with 4- to 6-week streptozotocin-induced diabetes. Electrically paced papillary muscles from diabetic rats produced contracture when exposed to the high concentration of Ca^2+ (～10 mM), whereas normal muscles exhibited a positive inotropic response to Ca^2+ with little elevation of resting tension. Measurement of intracellular Ca^2+ concentration (Ca^2+ _i ) using Indo-1 revealed that diastolic levels of Ca^2+ _i were increased in both myocytes from control and diabetic rats with increasing Ca^2+ _o from 1.3 to 5.2 mM. However, the increase in diastolic Ca^2+ _i was more marked in diabetic myocytes. In contrast, the increase in peak systolic Ca^2+ _i produced by elevated Ca^2+ _o was significantly higher in control myocytes. After the rapid application of caffeine (10 s after cessation of electrical stimulation) induced a large increase in Ca^2+ _i by releasing Ca^2+ from the sarcoplasmic reticulum, the decay of the Ca^2+ _i transient was slower in diabetic myocytes compared to controls. These results provide evidence that the mechanism underlying extrusion of Ca^2+ from myocytes, majorly the Na^+ /Ca^2+ exchanger, is impaired by diabetes.
We have previously reported that the positive inotropic response to β-adrenoceptor stimulation was markedly diminished in diabetic rat papillary muscles. This finding prompted us to characterize the ...properties of cardiac β-adrenoceptrors in diabetes by means of radioligand binding with ICYP. Male Wistar rats received 45 mg/kg streptozotocin to induce diabetes , and after a lapse of 4 weeks ventricular membrane fractions were prepared. Binding of ICYP to control and diabetic membranes was a saturable process of high affinity with very low non-specific binding. Scatchard analysis resulted in linear plots for both groups (K_D ：60-100 pM), indicating interaction with a single class of binding sites. The maximal number of ICYP binding sites, however, was about 40% less in diabetic group. The isoproterenol (ISO) displacement curve for ICYP showed two classes binding sites that have high and low affinity states for ISO. The proportion and dissociation constant of high affinity states were not different between both groups. GppNHp converted the heterogenous binding into a homogenous one of low affinity. If β-adrenoceptors expressing high, GppNHp-sensitive affinity for ISO have functionally an important role, their decrease under diabetes may explain in part the diminished inotropic response via β-adrenoceptors.
The antimuscarinic properties of pirmenol(Pr) and penticainide(Pt), new class Ia antiarrhythmic agents, were compared with those of disopyramide(D) in guinea pig left atrium(LA), urinary bladder(Bl) ...and submandibular gland(Gl). Pr, Pt and D all caused concentration-dependent rightward shift of the concentration-response curves for the carbachol-induced negative inotropic effect in LA and contraction in Bl. The antimuscarinic potencies of D and Pt in LA were similar to those in Bl. However, Pr was ten times more potent in LA than in Bl. The orders of antimuscarinic potency were Pr>D>Pt in LA and D>Pr>Pt in Bl. All the agents inhibited specific ^^3 H-N-methylscopolamine(NMS) binding to membranes from LA, Bl and Gl. Computer-assisted analysis of the displacement curves by Pr showed that Pr binds to one site in both LA and Gl. The Ki value for Pr in Gl was ten times greater than that in LA. In Bl, Pr was found to bind to two distinct affinity sites. The K_H and K_L values in Bl were comparable to the Ki values obtained in LA and Gl, respectively. On the other hand, both D and Pt inhibited ^^3 HNMS binding in LA in a manner analogous to that observed in Bl. In conclusion, while D and Pt does not exhibit selective affinity for muscarinic receptor subtypes, Pr may discriminate between cardiac and glandular subtypes of muscarinic receptors.
The antimuscarinic properties of pirmenol, a new class Ia antiarrhythmic agent, were compared with those of quinidine and disopyramide. In electrically driven left atria and spontaneously beating ...right atria of guinea-pigs, Pirmenol, quinidine and disopyramide all caused a concentration-dependent rightward shift of the concentration-response curves for the negative inotropic and chronotropic effects of carbachol. The order of the antimuscarinic potency was pirmenol>disopyramide>quinidine. The (3H)-N-methylscopolamine (NMS) binding to the membrane preparation from guinea-pig left atria was saturable with KD of 168 pM and Bmax of 284 fmol/mg of protein. All of the agents inhibited specific (3H)NMS binding in a concentration-dependent manner. The rank order of potency was identical to that determined by the functional study. At low concentrations, these agents produced dissociation of (3H)NMS, but only pirmenol dissociated (3H)NMS from the binding sites by an extent comparable to that of 1 uM atropine. On the other hand, at high concentrations, the agents including pirmenol strongly inhibited the dissociation of (3H)NMS elicited by 1 uM atropine. The results indicate that pirmenol, disopyramide as well as quinidine exert their antimuscarinic action possibly by interacting both competitively and allosterically with cardiac muscarinic receptors, although the mode by which pirmenol competes for (3H)NMS binding sites is different from those of others.
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