Comparisons of chromosome Y-substituted mouse strains reveal that the male-specific chromosome modulates the effects of androgens on cardiac functions

BACKGROUND: The C57BL/6J.Y(A/J) mouse strain is a chromosome-substituted line where the original male-specific portion of chromosome Y (MSY) from C57BL/6J mice was substituted for that from A/J mice. In hearts from male C57BL/6J.Y(A/J) and C57BL/6J mice, orchidectomy (ORX) affected in a strictly strain-specific fashion the expression a subset of genes showing enrichment for functional categories, including that of circadian rhythms and cardiac contractility. We further tested whether: (1) there were strain-specific differences in cardiac circadian rhythms; (2) strain-dependent differences in the effects of ORX on contractility genes translated into differences in cardiac functions; and (3) differential contractility responses occurred preferentially at times when circadian rhythms also showed strain-specific differences. METHODS: In hearts from the two above strains, we (1) profiled the expression levels of 15 circadian genes at 4-h intervals across a 24 h period; (2) tested the effects of either ORX or androgen replacement on expression of cardiac contractility genes, and that of ORX on myocardial functional reserve; and (3) verified whether the effects of MSY variants on cardiac contractility-related responses showed synchronicity with differences in circadian rhythms. RESULTS: Among the 15 tested circadian genes, a subset of them were affected by strain (and thus the genetic origin of MSY), which interacted with the amplitude of their peak of maximal expression at 2:00 PM. At that same time-point, ORX decreased (and androgen supplementation increased) the expression of three contractility-related genes, and decreased myocardial relaxation reserve in C57BL/6J.Y(A/J), but not in C57BL/6J mice. These effects were not detected at 10:00 AM, i.e., at another time-point when circadian genes showed no strain-specific differences. CONCLUSIONS: The results indicate that in mice, androgens have activational effects on cardiac circadian rhythms, contractile gene expression, and myocardial functional reserve. All effects occurred preferentially at the same time of the day, but varied as a function of the genetic origin of MSY. Androgens may therefore be necessary but not sufficient to impart male-specific characteristics to some particular cardiac functions, with genetic material from MSY being one other necessary factor to fully define their range of actions

Cyclic GMP inhibits a pharmacologically distinct Na+/H+ exchanger variant in cultured rat astrocytes via an extracellular site of action

There is growing evidence that cyclic GMP (cGMP) plays important roles in the brain. In cultured rat astrocytes, we observed that the cGMP-inducing C-type natriuretic peptide (CNP) and cGMP analogues caused a decrease in intracellular pH (pHi). To examine whether this effect was due to inhibition of an Na+/H+ exchanger (NHE), we acidified cells by replacing extracellular Na+ by choline and examined the kinetics of the pHi recovery that occurred on reintroduction of Na+ in the extracellular medium. Both CNP and amiloride analogues inhibited the Na+-dependent pHi recovery, even in the nominal absence of CO2/HCO3−. This indicated that CNP inhibited the activity of an exchanger that was Na+-dependent, HCO3−-independent, and sensitive to known inhibitors of NHE. However, comparison of the potencies of four distinct amiloride analogues revealed a pharmacological profile that was different from that of any other NHE characterized to date. cGMP mimicked the effect of CNP on sodium-dependent pHi recovery, but the native nucleotide was as potent as membrane-permeant analogues. Intracellularly produced cGMP was very rapidly exported out of astrocytes. Probenecid and niflumic acid slowed down the rate of cGMP egression and inhibited the effect of CNP on Na+-dependent recovery, but not that of extracellular cGMP. Altogether, our data indicate that cGMP inhibits a novel type of NHE in astrocytes via an extracellular site of action. If these results with primary cultures transfer to brain, this phenomenon may constitute a mechanism by which natriuretic peptides exert some of their actions in the brain, as pHi transients have been shown to modulate several important astrocytic functions

Characterization of myocardium, isolated cardiomyocytes, and blood pressure in WKHA and WKY rats

We previously reported that the left ventricular (LV) mass of Wistar-Kyoto (WKY)-derived hyperactive (WKHA) rats was higher than that of WKY rats in the absence of a difference in systolic blood pressure. To extend these earlier observations, we conducted a series of functional and morphological investigations on both strains. Analysis of tissue sections revealed that the surface of ventricular tissue from WKHA rats was higher than that of WKY rats, without any enlargement of the cavity area. Analysis of isolated adult cells showed that cell width (as well as cell volume) of ventricular cardiomyocytes was significantly higher in WKHA than WKY rats. However, LV of WKHA rats contained approximately 33% less cardiomyocytes than those from WKY rats. Mean intracellular free calcium concentration of cardiomyocytes was also higher in WKHA than WKY rats. Hemodynamic measurements revealed that the values of the maximum rates of pressure change (dP/dt) were higher in LV from WKHA rats. However, these differences were reduced (-dP/dt) or abolished (+dP/dt) when the values were normalized for both the number and mean cross-sectional area of ventricular cardiomyocytes. Mean levels of systolic and diastolic blood pressure (corresponding to the 24-h average of measurements obtained continuously in conscious unrestrained animals using radiotelemetric implants) were not different between strains. However, circadian rhythm was more evident in WKY rats, because the difference between morning and night values of systolic and diastolic blood pressure was greater (by 3 mmHg) in WKY rats. Altogether, our data validate the use of WKHA rats as models of predominantly concentric LV hypertrophy developing in the absence of increased mean levels of hemodynamic cardiac load and show that the hypertrophy phenotype is more pronounced in isolated cardiomyocytes than at the level of the whole ventricle

Differences in cell-type-specific responses to angiotensin II explain cardiac remodeling differences in C57BL/6 mouse substrains

Despite indications that hearts from the C57BL/6N and C57BL/6J mouse substrains differ in terms of their contractility and their responses to stress-induced overload, no information is available about the underlying molecular and cellular mechanisms. We tested whether subacute (48 hours) and chronic (14 days) administration of angiotensin II (500 ng/kg per day) had different effects on the left ventricles of male C57BL/6J and C57BL/6N mice. Despite higher blood pressure in C57BL/6J mice, chronic angiotensin II induced fibrosis and increased the left ventricular weight/body weight ratio and cardiac expression of markers of left ventricular hypertrophy to a greater extent in C57BL/6N mice. Subacute angiotensin II affected a greater number of cardiac genes in C57BL/6N than in C57BL/6J mice. Some of the most prominent differences were observed for markers of (1) macrophage activation and M2 polarization, including 2 genes (osteopontin and galectin-3) whose inactivation was reported as sufficient to prevent angiotensin II-induced myocardial fibrosis; and (2) fibroblast activation. These differences were confirmed in macrophage- and fibroblast-enriched populations of cells isolated from the hearts of experimental mice. When testing F2 animals, the amount of connective tissue present after chronic angiotensin II administration did not cosegregate with the inactivation mutation of the nicotinamide nucleotide transhydrogenase gene from C57BL/6J mice, thus discounting its possible contribution to differences in cardiac remodeling. However, expression levels of osteopontin and galectin-3 were cosegregated in hearts from angiotensin II-treated F2 animals and may represent endophenotypes that could facilitate the identification of genetic regulators of the cardiac fibrogenic response to angiotensin II