Yoshihisa Mio

Nitric oxide-dependent vasodilator mechanism is not impaired by hypertension but is diminished with aging in the rat aorta.

This study was designed to elucidate the effects of hypertension and aging on nitric oxide (NO)-mediated relaxation response to acetylcholine in the rat aorta. NO-mediated relaxation response was assessed as the relaxation response to acetylcholine after treatment with cyclooxygenase inhibitor in KCl-precontracted aortic rings. The endothelium-dependent relaxation responses to acetylcholine were lower in aortic rings isolated from spontaneously hypertensive rats (SHRs) at ages 16-20 and 55-60 weeks compared with those seen in age-matched Wistar-Kyoto (WKY) rats. Aging induced a reduction of the relaxation response to acetylcholine in aortic rings from WKY rats but not from SHRs. Pretreatment with indomethacin enhanced the relaxation response to acetylcholine in only SHRs at ages 16-20 and 55-60 weeks, thereby cancelling the difference in the relaxation response between WKY rats and SHRs. Simultaneous administration of indomethacin and NG-nitro-L-arginine methyl ester abolished the relaxation response to acetylcholine in both strains. Thus NO-mediated relaxation response to acetylcholine was similar between WKY rats and SHRs at ages 16-20 and 55-60 weeks, respectively, and was attenuated with aging to the same degree in both strains. In conclusion, NO-mediated relaxation response to acetylcholine in the aorta is attenuated with aging but not impaired by hypertension.

Ca2+ entry through the store-mediated pathway directly activates only the K+ current but the subsequent Ca2+ release from the store activates both K+ and Cl− currents in submandibular gland acinar cells of the rat

The store-mediated Ca2+ entry was detected in single and cluster of rat submandibular acinar cells by measuring the Ca2+ activated ionic membrane currents. In the cells where intracellular Ca2+ was partly depleted by stimulation with submaximal concentration of acetylcholine (ACh) under a Ca2(+)-free extracellular condition, an employment of external Ca2+ in the absence of ACh caused a sustained increase of the K+ current without affecting the Cl- current. A renewed ACh challenge without external Ca2+ caused repetitive spikes of both K+ and Cl- currents due to the Ca2+ release. SK & F 96365 inhibited the generation of the sustained K+ current and refilling of the Ca2+ store following the Ca2+ readmission. It is suggested that the Ca2+ enters the cell through the store-mediated pathway new the K+ channels and is taken up by the store. Thus, only Ca2+ released from the store can activate both the K+ and Cl- currents.

Oxidized LDLs but not native LDLs augment Ba2+ currents through L-type Ca2+ channels of the A7r5 smooth muscle-derived cell line.

The whole-cell patch-clamp method was used on A7r5 smooth muscle-derived cell line, and Ba2+ currents through Ca2+ channels were recorded. The A7r5 cells showed voltage-dependent, long-lasting Ba2+ currents which were markedly inhibited by nifedipine (10 microM). The magnitude of the maximum Ba2+ current (IBa(max)) was augmented by an application of dbcAMP (1 mM), but not affected by TPA (80 nM). Noradrenaline (NA) at 100 microM caused an increase in the IBa(max) by 19.7% in the presence of phentolamine (10 microM). This effect was cancelled by Rp-cAMPs (10 microM). In the presence of propranolol (10 microM), NA tended to reduce the IBa(max). Application of Ox-LDLs at 100 microg protein/ml caused an increase in the IBa(max) by 15.7%, whereas native LDLs did not change the IBa(max). Rp-cAMPs was ineffective to the Ox-LDL action on the IBa(max). In the presence of Ox-LDLs, NA augmented the IBa(max) by 21.4% in the presence of phentolamine. These results suggest that Ox-LDLs activate L-type Ca2+ channels of A7r5 cells by a mechanism independent of cAMP/PKA signalling.

Excitatory and inhibitory actions of norepinephrine on the Ba2+ current through L-type Ca2+ channels of smooth muscle cells of guinea-pig vas deferens.

The effect of norepinephrine (NE) was examined on the whole‐cell Ba2+ current through L‐type Ca2+ channels of freshly isolated smooth muscle cells of guinea‐pig vas deferens. The magnitude of maximum Ba2+ current [IBa(max)] varied in different cells, although the capacitance of the cell membrane was similar (∼50 pF). Application of dbcAMP augmented IBa(max) by 37%, which was canceled by Rp‐cAMPs, while PMA decreased the current by 32%, which was canceled by staurosporine. NE increased IBa(max) of the cells which originally showed relatively small IBa(max), and decreased the current of the cells which showed larger IBa(max). In the presence of phentolamine, NE increased IBa(max), and this effect was remarkable in cells showed smaller IBa(max). In the presence of propranolol, NE decreased IBa(max). The excitatory β‐adrenoceptor activation was canceled by Rp‐cAMPs, and the inhibitory α‐adrenoceptor effect was canceled by staurosporine. It is suggested that NE shows dual (excitatory and inhibitory) actions on the L‐type Ca2+ channels of smooth muscle of guinea‐pig vas deferens. The excitatory β‐adrenoceptor action mediated through cAMP/PKA is predominant in cells with lower density of the Ca2+ channels, while inhibitory α‐adrenoceptor action mediated through PKC is predominant in cells with higher channel density.

Role of extracellular Ca2+ in acetylcholine‐induced repetitive Ca2+ release in submandibular gland acinar cells of the rat

Acetylcholine (ACh) caused repetitive transient Cl− currents activated by intracellular Ca2+ in single rat submandibular grand acinar cells. As the concentration of ACh increased the amplitude and the frequency of the transient Cl− currents increased. These responses occurred also in the absence of extracellular Ca2+ but disappeared after several minutes. Repetitive transient Cl− currents were restored by readmission of Ca2+ to the extracellular solution. The higher the concentration of extracellular Ca2+ readmitted, the larger the amplitude of the transient Cl− currents. Ca2+ entry through a store‐coupled pathway was detected by application of Ca2+ to the extracellular solution during a brief cessation of stimulation with ACh. In these experiments too, the higher the concentration of Ca2+, the larger the transient Cl− currents activated by Ca2+ released from the stores. The time course of decrease in total charge movements of repetitive transient responses to ACh with removal of extracellular Ca2+ depended on a decrease in charge movements of each transient event rather than a decrease in frequency of the repetitive events. The decrease of charge movements of each transient event was due to a decrease in its amplitude rather than its duration. The results suggest that in this cell type an amplitude‐modulated mechanism is involved in repetitive Ca2+ release and that Ca2+ entry is essential to maintain the repetitive release of Ca2+. The results further suggest that the magnitude of Ca2+ entry determines the number of unitary stores filled with Ca2+ which can synchronously respond to ACh.