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The Na + -HCO 3 − cotransporter (NBC) is an important sarcolemmal acid extruder in cardiac muscle. The characteristics of NBC expressed functionally in heart are controversial, with reports suggesting electroneutral (NBCn; 1HCO 3 − : 1Na + ; coupling coefficient N = 1) or electrogenic forms of the transporter (NBCe; equivalent to 2HCO 3 − : 1Na + ; N = 2). We have used voltage-clamp and epifluorescence techniques to compare NBC activity in isolated ventricular myocytes from rabbit, rat and guinea pig. Depolarization (by voltage clamp or hyperkalaemia) reversibly increased steady-state pH i while hyperpolarization decreased it, effects seen only in CO 2 /HCO 3 − -buffered solutions, and blocked by S0859 (cardiac NBC inhibitor). Species differences in amplitude of these pH i changes were rat > guinea pig ≈ rabbit. Tonic depolarization (−140 mV to −0 mV) accelerated NBC-mediated pH i recovery from an intracellular acid load. At 0 mV, NBC-mediated outward current at resting pH i was +0.52 ± 0.05 pA pF −1 (rat, n = 5), +0.26 ± 0.05 pA pF −1 (guinea pig, n = 5) and +0.10 ± 0.03 pA pF −1 (rabbit, n = 9), with reversal potentials near −100 mV, consistent with N = 2. The above results indicate a functionally active voltage-sensitive NBCe in these species. Voltage-clamp hyperpolarization negative to the reversal potential for NBCe failed, however, to terminate or reverse NBC-mediated pH i -recovery from an acid load although it was slowed significantly, suggesting electroneutral NBC may also be operational. NBC-mediated pH i recovery was associated with a rise of [Na + ] i at a rate ∼25% of that mediated via NHE, and consistent with an apparent NBC stoichiometry between N = 1 and N = 2. In conclusion, NBCe in the ventricular myocyte displays considerable functional variation among the three species tested (greatest in rat, least in rabbit) and may coexist with some NBCn activity.

Endothelin-1 (ET-1) has been implicated in fibroblast proliferation. However, the mechanism involving ET-1 is not clear. The present study was performed to examine the role of endogenous ET-1 in ET-1-stimulated fibroblast proliferation and to investigate the regulatory mechanism of ET-1-induced ET-1 gene expression in cardiac fibroblasts. Both ET A receptor antagonistand endothelin-converting enzyme inhibitor (phosphoramidon) inhibited the increased DNA synthesis caused by ET-1. ET-1 gene was induced by ET-1, as revealed with Northern blotting and ET-1 promoter activity assay. ET-1 increased intracellular reactive oxygen species (ROS), which were significantly inhibited by BQ485 and antioxidants. Antioxidants suppressed ET-1 gene expression and DNA synthesis stimulated by ET-1. ET-1 activated mitogen-activated protein kinases (MAPK), including extracellular signal-regulated kinase (ERK), p38 MAPK, and c-Jun N-terminal kinase, which were significantly inhibited by antioxidants. Only ERK inhibitor U0126 could inhibit ET-1-induced transcription of the ET-1 gene. Cotransfection of dominant-negative mutant of Ras, Raf, and MEK1 decreased the ET-1-induced increase in ET-1 transcription, suggesting that the Ras-Raf-ERK pathway is required for ET-1 action. Truncation and mutational analysis of the ET-1 gene promoter showed that the activator protein-1 (AP-1) binding site was an important cis-element in ET-1-induced ET-1 gene expression. Antioxidants attenuated the ET-1-stimulated AP-1 binding activity. Our data suggest that ROS were involved in ET-1-induced fibroblast proliferation and mediated ET-1-induced activation of ERK pathways, which culminated in ET-1 gene expression.

1 Our previous study demonstrated that the aortic inducible nitric oxide synthase (iNOS) expression and the plasma nitrite level in spontaneously hypertensive rats (SHR) were greater than that in age-matched Wistar-Kyoto rats (WKY). We subsequently hypothesized that the overexpression of iNOS might play an important role in the pathogenesis of hypertension in SHR. 2 In the present study, pyrrolidinedithiocarbamate (PDTC, 10 mg kg 71 day 71 , p.o., antioxidant and nuclear factor-k B inhibitor) and aminoguanidine (15 mg kg 71 day 71 , p.o., selective inhibitor of iNOS) was used to treat SHR and WKY from age of 5 weeks through 16 weeks. 3 We found that PDTC and aminoguanidine signi®cantly suppressed the development of hypertension and improved the diminished vascular responses to acetylcholine in SHR but not in WKY. Likewise, the increase of iNOS expression, nitrotyrosine immunostaining, nitric oxide production and superoxide anion formation in adult SHR were also signi®cantly suppressed by chronic treatment with PDTC and aminoguanidine. 4 In conclusion, this study demonstrated that both PDTC and aminoguanidine signi®cantly attenuated the development of hypertension in SHR. The results suggest that PDTC suppresses iNOS expression due to its anti-oxidant and/or nuclear factor-k B inhibitory properties. However, the e ect of aminoguanidine was predominantly mediated by inhibition of iNOS activity, thereby reducing peroxynitrite formation. We propose that the development of a more speci®c and potent inhibitor of iNOS might be bene®cial in preventing pathological conditions such as the essential hypertension.

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