Effects of total replacement of atrial myosin light chain-2 with the ventricular isoform in atrial myocytes of transgenic mice.

BACKGROUND: In contrast to their well-known and critical role in excitation-contraction coupling of vascular smooth muscle, the effects of the myosin light chains on cardiomyocyte mechanics are poorly understood. Accordingly, we designed the present experiment to define the cardiac chamber-specific functional effects of the ventricular isoform of the regulatory myosin light chain (MLC2v). METHODS AND RESULTS: Postnatal transgenic cardiac-specific overexpression of MLC2v was achieved by use of the alpha-myosin heavy chain promoter. Enzymatically disaggregated atrial and ventricular mouse myocytes were field-stimulated at multiple frequencies, and mechanical properties and calcium kinetics were studied by use of video edge detection and FURA 2-AM, respectively. MLC2v overexpression resulted in complete replacement of the atrial with the ventricular isoform of the regulatory myosin light chain at the steady-state mRNA and protein levels in the atria of transgenic mice. Mechanical properties of transgenic atrial myocytes were enhanced to the level of ventricular myocytes of control animals in association with modest decreases in the amplitude of the calcium transient. CONCLUSIONS: MLC2v modulates chamber-specific contractility by enhanced calcium sensitivity and/or improved cross-bridge cycling of the thin and thick filaments of the cardiomyocyte.

The effects of a thyroid hormone analog on left ventricular performance and contractile and calcium cycling proteins in the baboon.

To determine the biochemical and related functional effects of the thyroid analog diiodothyroproprionic acid (DITPA) on primate myocardium, we examined, both before and after 23 days of DITPA (3.75 mg/kg): myosin heavy-chain (MHC) isoforms and sarcoplasmic reticulum (SR) calcium cycling proteins; left ventricular (LV) function; and the LV force-frequency relation in four baboons chronically instrumented with sonomicrometers and micromanometers. The force-frequency relation was measured as the response of isovolumic contraction (dP/dtmax) to incremental pacing and the critical heart rate (HRcrit) as the rate at which dP/dtmax reached its maximum. DITPA increased basal LV dPt/dtmax (3,300 +/- 378 versus 2,943 +/- 413 mm Hg/sec; p = .09), and velocity of circumferential shortening (1.13 +/- 0.30 versus 0.76 +/- 0.30 circ/sec; p < .01), decreased the basal time constant of isovolumic relaxation (24.2 +/- 1.6 versus 29.9 +/- 2.5 msec; p < .05), and increased the HRcrit (203 +/- 19 versus 168 +/- 20 bpm; p < .05), without effecting significant changes in either basal heart rate (119 +/- 14 versus 111 +/- 17 bpm) or systolic blood pressure (137 +/- 14 versus 126 +/- 8 mm Hg). Quantitative immunoblotting revealed significant decreases in both phospholamban and the ratio of phospholamban to SR Ca2+ adenosine triphosphatase in DITPA-treated animals when compared to four untreated controls. By contrast, alpha-MHC isoform was undetectable in both DITPA treated and control baboons. Thus, DITPA favorably alters the stoichiometry between the SR calcium pump and its inhibitor, phospholamban, and has positive inotropic and lusitropic effects in the normal primate left ventricle, which may be useful in the treatment of heart failure. Unlike thyroid hormone, these changes occur in the absence of detectable alpha-MHC isoform protein expression and without an increase in heart rate.

In vivo determination of left ventricular wall stress-shortening relationship in normal mice.

Although targeted alterations of the mouse genome are used increasingly to identify the mechanisms underlying cardiac function, the methods used to study the phenotypic expression of these alterations in vivo are limited. To derive a relatively noninvasive, load-independent measure of left ventricular (LV) contractility in mice, we cannulated the femoral artery and performed two-dimensional directed M-mode echo studies in 28 anesthetized FVB/N mice, using a 9-MHz transducer. Loading conditions were altered by intraarterial methoxamine (3-12 microg/g), and LV shortening fraction was determined at several steady states, both before and after myocardial contractility was altered by either 4 microg/g intraperitoneal dobutamine (n = 16) or 1-2 microg/g verapamil (n = 12). The relation between LV systolic meridional stress and fractional shortening derived from pooled baseline data was inverse and linear [r = 0.80, slope = -0.19, intercept = 48%, standard error of estimate (SEE) = 5.5%, P < 0.001]. Dobutamine produced a parallel upward shift of the relation (r = 0.87, slope = -0.21, intercept = 61%, SEE = 4.5%, P < 0.001), and verapamil produced a downward shift of the relation (r = 0.48, slope = -0.05, intercept = 24%, SEE = 3.7%, P < 0.05). At matched levels of end-systolic stress, dobutamine increased and verapamil decreased the LV shortening fraction. We conclude that 1) inverse stress-shortening relations can be assessed noninvasively in mice; and 2) these relations are sensitive to alterations in inotropic state, independent of loading conditions.

Effects of thyroid hormone on left ventricular performance and regulation of contractile and Ca(2+)-cycling proteins in the baboon. Implications for the force-frequency and relaxation-frequency relationships.

The transcriptional, posttranscriptional, and related functional effects of thyroid hormone on primate myocardium are poorly understood. Therefore, we studied the effects of thyroid hormone on sarcoplasmic reticulum (SR) Ca(2+)-cycling proteins and myosin heavy chain (MHC) composition at the steady state mRNA and protein level and associated alterations of left ventricular (LV) performance in 8 chronically instrumented baboons. The force-frequency and relaxation-frequency relations were assessed as the response of LV isovolumic contraction (dP/dtmax) and relaxation (Tau), respectively, to incremental atrial pacing. Both the heart rate at which dP/dtmax was maximal and Tau was minimal (critical heart rates) in response to pacing were increased significantly after thyroid hormone. Postmortem LV tissue from 5 thyroid-treated and 4 additional control baboons was assayed for steady state mRNA levels with cDNA probes to MHC isoforms and SR Ca(2+)-cycling proteins. Steady state SR Ca(2+)-ATPase and phospholamban mRNA increased in the hyperthyroid state, and alpha-MHC mRNA appeared de novo, whereas beta-MHC mRNA decreased. Western analysis (4 thyroid-treated and 4 control baboons) showed directionally similar changes in MHC isoforms and a slight increase in SR Ca(2+)-ATPase. In contrast, there was a statistically nonsignificant decrease in phospholamban protein, which resulted in a significant 40% decrease in the ratio of phospholamban to SR Ca(2+)-ATPase. Thus, thyroid hormone increases the transcription of Ca(2+)-cycling proteins and shifts MHC isoform expression in the primate LV. Our data suggest that both transcriptional and posttranslational mechanisms determine the levels of these proteins in the hyperthyroid primate heart and mediate, in part, the observed enhanced basal and frequency-dependent LV performance.

Increased dietary salt sensitizes vasomotor neurons of the rostral ventrolateral medulla.

Excess dietary sodium is a major contributing factor to the incidence and severity of hypertension. However, the precise mechanism or mechanisms by which salt contributes to the severity of hypertension are unknown. The region of the rostral ventrolateral medulla (RVLM) is a principal brain stem locus critical for the regulation of arterial blood pressure by the sympathetic nervous system. The purpose of this study was to determine if excess dietary sodium chloride might alter the function or responsiveness of neurons in the RVLM. Male Sprague-Dawley rats were given either tap water or 0.9% sodium chloride solution to drink for 10 to 14 days. Excess sodium chloride did not affect baseline blood pressure. However, when neurons of the RVLM were stimulated by microinjections of L-glutamate, evoked increases in arterial pressure were potentiated in rats given sodium chloride. Augmented pressor responses could not be accounted for by increased vascular reactivity because both groups responded similarly to intravenously administered phenylephrine and norepinephrine. Additionally, electrical stimulation of descending spinal sympathoexcitatory axons produced identical pressor responses in both groups, indicating that altered synaptic transmission at central or peripheral neuroeffector junctions distal to the RVLM could not explain enhanced pressor responses produced by direct stimulation of RVLM cell somata. Finally, impaired arterial baroreceptor reflexes could not account for augmented RVLM pressor responses, as depressor and bradycardic responses produced by electrical stimulation of aortic baroreceptor afferents were not reduced in rats given excess dietary sodium chloride.(ABSTRACT TRUNCATED AT 250 WORDS)