Volume overload cardiac hypertrophy exhibits decreased expression of g(s)alpha and not of g(i)alpha in heart.

We have recently reported enhanced levels of G(i)alpha proteins in genetic and other experimentally induced models of hypertension, whereas the levels of G(s)alpha were decreased in hypertensive rats expressing cardiac hypertrophy. The present studies were undertaken to investigate whether the decreased levels of G(s)alpha are associated with cardiac hypertrophy per se and used an aortocaval fistula (AV shunt; volume overload) rat model that exclusively expresses cardiac hypertrophy. Cardiac hypertrophy in Sprague-Dawley rats (200-250 g) was induced under anesthesia, and, after a period of 10 days, the hearts were used for adenylyl cyclase activity determination, protein quantification, and mRNA level determination. A temporal relationship between the expression of G(s)alpha proteins and cardiac hypertrophy was also examined on days 2, 3, 7, and 10 after induction of AV shunt in the rat. The heart-to-body-weight ratio (mg/g) was significantly increased in AV shunt rats after 3, 7, and 10 days of induction of AV shunt compared with sham-operated controls, whereas arterial blood pressure was not different between the two groups. Guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS) stimulated adenylyl cyclase activity in a concentration-dependent manner in heart membranes from both groups; however, the degree of stimulation was significantly decreased in AV shunt rats. In addition, the stimulatory effects of isoproterenol were also diminished in AV shunt rats compared with control rats, whereas glucagon-stimulated adenylyl cyclase activity was not different in the two groups. The inhibitory effects of oxotremorine (receptor-dependent G(i) functions) and low concentrations of GTPgammaS on forskolin-stimulated adenylyl cyclase activity (receptor-independent G(i) functions) were not different in the two groups. In addition forskolin and NaF also stimulated adenylyl cyclase activity to a lesser degree in AV shunt rats compared with control rats. The levels of G(i)alpha-2 and G(i)alpha-3 proteins and mRNA, as determined by immunoblotting and Northern blotting, respectively, were not different in both groups; however, the levels of G(s)alpha(45) and G(s)alpha(47), and not of G(s)alpha(52), proteins were significantly decreased in AV shunt rats by days 7 and 10 compared with control rats, whereas no change was observed on days 2 and 3 after induction of AV shunt. These results suggest that the decreased expression of G(s)alpha proteins may not be the cause but the effect of hypertrophy and that the diminished responsiveness of adenylyl cyclase to GTPgammaS, isoproterenol, NaF, and forskolin in hearts from AV shunt rats may partly be due to the decreased expression of G(s)alpha. It can be concluded from these studies that the decreased expression of G(s)alpha may be associated with cardiac hypertrophy and not with arterial hypertension.

Enhanced expression of Gi proteins in non-hypertrophic hearts from rats with hypertension-induced by L-NAME treatment.

OBJECTIVE: The objective of the present studies is to investigate if the enhanced expression of Gs alpha protein and their mRNA observed in various models of hypertensive rats is due to the expressed hypertrophy or hypertension. METHODS: Hypertension, in Sprague-Dawley rats was induced by the oral administration of the arginine analog N(omega)-nitro-L-arginine methyl ester (L-NAME) in their drinking tap water for a period of 4 weeks. The control rats were given plain tap water only. The levels of inhibitory guanine nucleotide regulatory proteins (Gi alpha-2, Gi alpha-3), stimulatory guanine nucleotide proteins (Gs alpha) and G beta proteins were determined by immunoblotting, whereas the levels of Gi alpha-2, Gi alpha-3, Gs alpha and adenylyl cyclase type V enzyme mRNA were determined by Northern-blotting techniques. Adenylyl cyclase activity was determined by measuring [32P]cAMP formation from [alpha32P]ATP. RESULTS: The systolic blood pressure was enhanced in L-NAME-treated rats compared to control rats (190 +/- 9.2 mmHg versus 121 +/- 6.3 mmHg); however, heart-to-body-weight ratio was not different in two groups. The levels of Gi alpha-2 and Gi alpha-3 proteins and their mRNA were significantly augmented in hearts from L-NAME-treated rats, however, the levels of Gs alpha and G beta were unaltered. In addition, the effect of low concentrations of GTPgammaS on forskolin (FSK)-stimulated adenylyl cyclase activity (receptor-independent functions of Gi alpha) was significantly enhanced in L-NAME-treated rats. However, the inhibitions of adenylyl cyclase exerted by oxotremorine, C-ANP(4-23) and angiotensin II (AII) (receptor-dependent function of Gi alpha) were completely attenuated in L-NAME-treated rats. On the other hand, cholera toxin stimulated GTP or GTPgammaS-sensitive adenylyl cyclase activity (Gs alpha function) to similar extent in control and L-NAME-treated rats, suggesting that Gs alpha functions were not altered by L-NAME treatment. However, the stimulatory effects of isoproterenol, glucagon, NaF on adenylyl cyclase were diminished in L-NAME-treated rats. In addition, FSK-stimulated enzyme activity was also diminished in L-NAME-treated rats without any changes in the mRNA levels of type V enzyme. CONCLUSIONS: These results suggest that L-NAME hypertensive rats that do not express cardiac hypertrophy exhibit enhanced expression of Gi alpha protein and associated adenylyl cyclase activity.

Nitric oxide synthase inhibition by N(omega)-nitro-L-arginine methyl ester modulates G-protein expression and adenylyl cyclase activity in rat heart.

We have recently shown an enhanced expression of inhibitory guanine nucleotide regulatory proteins Gi alpha-2 and Gi alpha-3 and their respective mRNA in hearts from DOCA-salt hypertensive rats. However, it is not known whether these changes are due to the expressed hypertrophy or hypertension. The present studies were therefore undertaken to investigate this possibility. Hypertension in Sprague-Dawley rats was induced by the oral administration of the arginine analog N(omega)-nitro-L-arginine methyl ester (L-NAME) in their drinking tap water for a period of 4 weeks. The control rats were given plain tap water only. L-NAME-treated rats showed an enhanced blood pressure (190 +/- 9.23 mm Hg; n = 20) compared to control rats (121 +/- 6.3 mm Hg; n = 20). However, heart to body weight ratio was not different in the two groups. Guanosine 5'-o-(3-thiotriphosphate) (GTPgammaS) stimulated adenylyl cyclase activity in heart membranes from both groups, but the extent of stimulation was significantly decreased in L-NAME-treated rats. Similarly, stimulations exerted by isoproterenol, glucagon, NaF, and forskolin on adenylyl cyclase were also diminished in L-NAME-treated rats. On the other hand, the inhibitory effect of low concentrations of GTPgammaS on forskolin-stimulated enzyme activity was significantly enhanced. The extent of oxotremorine-mediated inhibition of adenylyl cyclase was unaltered in both control and L-NAME-induced hypertensive rats. The levels of Gi alpha-2 and Gi alpha-3, but not of stimulatory guanine nucleotide regulatory protein Gs alpha, as determined by immunoblotting, were significantly augmented in L-NAME-treated rats. Northern blot studies revealed a significant increase in Gi alpha-2 and Gi alpha-3 mRNA with no changes in Gs alpha mRNA. These results suggest that the altered expression of Gi alpha proteins and adenylyl cyclase activity in L-NAME-treated rats may be attributed to hypertension and not to hypertrophy.