Correlation between biochemical properties and adaptive diversity of skeletal muscle myofibrils and myosin of some air-breathing teleosts.

Functional properties of myofibrils and relative stability of myosin of five teleosts Channa punctata, Clarias batrachus, M astacembalus armatus, Labeo rohita and Catla catla adapted to different breathing modes were compared. Myofibrillar contractility and m-ATPase of air-breathing organ (ABO) possessing C.punctata and C. batrachus were low and least affected by pH in the range of 7.1-8.5. However, their myosin isoforms were relatively thermostable, more soluble at sub-neutral pH values, between 0.1 to 0.15 M KCl concentrations and less susceptible to a-chymotryptic digestion. In contrast, myofibrils and myosin of water-breather major carps L. rohita and C. catla were more contractile and susceptible to pH and salt concentrations. Thus, correlation between catalytic efficiency and relative stability of myofibrils and myosin of ABO-possessing teleosts was of reverse order and magnitude, as compared to water-breathers. Interestingly, myofibrils and myosin of the behavioral air-breather M. armnatus showed intermediate properties. The specific levels of m-ATPase of all the five teleosts were in conformity with the levels of metabolic marker, the lactate dehydrogenase. The effect of chymotryptic cleavage of 94 and 173 kDa domains on ATPase, individuality of peptide maps of MyHC isomers and perturbation of phenylalanine residues by urea implicated hydrophobic residues in stabilizing myosin structure in these fish. The present study suggests two apparent evolutionary modifications of myofibrils and myosin in ABO-possessing teleosts: (i), 'down-regulation' of ATPase that explains sluggishness of such species and, (ii), more stable molecular structure to support stress of air-breathing modes of life.

Effect of caffeine on the Ca2+ pool affecting contractility and actomyosin ATPase activity in vascular smooth muscle of rabbit

Caffeine has been known to induce the contraction of rabbit aortic ring resulting from Ca2+ release from the intracellular stores. But in contrast, contraction of aortic ring induced by depolarizing agents or agonist was reported to be suppressed by caffeine. The present study was intended to examine the effect of caffeine on Ca2+ movement across the plasma membrane and actomyosin ATPase activity of vascular smooth muscle to elucidate the modes of action of caffeine on the vascular smooth muscle. Aortic ring preparation were made from the rabbit thoracic aorta and the endothelial cells were removed from the ring by gentle rubbing. The contractilty of the aortic ring was measured under varying conditions, and Ca2+ influx across the membranes of the aortic ring was measured with Ca2+ sensitive electrode with and without caffeine and the effect of caffeine on actomyosin ATPase activity were measured by modified Hartshrone's method. 45Ca wash out curves with and without caffeine were studied by Richard's method. The results were summarized as follows: 1) Caffeine inhibited the contractilty induced by norepinephrine. high K+, and histamine. but caffeine alone induced a transient contraction of vascular smooth muscle. The caffeine induced contraction was demonstrable even in the absence of external Ca2+. 2) Caffeine increased 45Ca efflux from vascular smooth muscle. 3) In the presence of propranolol, the inhibitory effect of caffeine on epinephrine induced contraction still persisted. 4) Caffeine decreased norepinephrine induced Ca2+ influx through the plasma membranes of aortic ring. 5) Caffeine decreased the actomyosin ATPase activity of vascular smooth muscle. From the above results, it is suggested that caffeine induces the contraction of vascular smooth muscle by release of Ca2+ from intracellular Ca2+ stone, but inhibits drug-induced contraction by decrease of Ca2+ influx across the plasma membranes and a decreased Ca2+ sensitivity of contractile protein in vascular smooth muscle.

A Study on the Cardiac Myofibrillar ATPase Activity in Diabetic Rats

Diabetes mellitus is known to be associated with a specific cardiomyopathy. This is evident from the clinical-pathological work and the epidemiologic data. An investigation was made in this study to determine whether diabetic cardiomyopathy in rats is associated with an alteration of biochemical characteristics of cardiac contractile proteins. Rats were made diabetic with intravenous injection of streptozotocin and hearts removed 8 weeks later for the isolation of myofibrils. The basal ATPase activity of myofibrils from diabetic hearts was significantly lower than that of the controls, suggesting the presence of some subtle structural and conformational changes in diabetic myofibrils. The activating effect of Mg ions on the myofibrillar actomyosin system of rat heart muscle was also demonstrated. Sodium dodecylsulfate gel electrophoresis showed the presence of myosin heavy chain, light chain 1 and 2, actin and troponin but failed to reveal differences in the patterns of these contractile proteins of light subunits between diabetics and controls. The deficiency in utilization of energy rich phosphates by the myofibrillar protein may be one of of the main mechanisms of cardiodepression observed in diabetic hearts. The cardiac myofibrillar ATPase activity may be one of useful measurements in evaluating pathophysiological states of cardiac contractile proteins.

pH-Temperature Dependence of Actomyosin Superprecipitation

The effect of temperature on the pH-dependence of actomyosin superprecipitation was studied, using actomyosin extracted from the rabbit and frog skeletal muscle tissues. The pH optima of superprecipitation was rather broad in both the rabbit and frog actomyosin. In the frog, superprecipitation measured at 16-42 degrees C was relatively independent of pH variations between 6.7 to 8.5, but it was significantly inhibited at pHs outside of this range, showing a sharp inflection of the curve. The pH at the inflection point was inversely proportional to the incubation temperature, but the (OH-)/(H+) ratio at the inflection point was not changed with temperature. The log (OH-)/(H+) was approximately -0.6 on the acidic side and 3.16 on the alkaline side. Similarly, superprecipitation of the frog actomyosin was virtually independent of the medium pH of the intermediate range (approximately 6.0-8.5); but it was drastically inhibited at pHs below or above this range, thus revealing a sharp inflection of the curve. Again, the pH at the inflection point changed inversely with temperature, preserving a constant (OH-)/(H+) ratio. The log (OH-)/(H+) ratio at the inflection point was approximately -2 on the acidic side and 3.5 on the alkaline side. The above pH effects were not associated with irreversible protein damage or with the changes in buffer species. These results strongly suggest that suppression of the superprecipitation of rabbit and frog actomyosin gels, at a low and high pH, be due to alterations in the fractional dissociation of histidine-imidazole and cysteine-SH groups, respectively.