Chronic salt-loading downregulates large-conductance Ca^(2+)-activated potassium channel in mesenteric arterial smooth muscle cells from SD rats

Objective Large-conductance calcium-activated potassium(BKCa)channel modulates vascular smooth muscle tone.In the present study,we tested the hypothesis that salt,one of the factors which significantly influence blood pressure(BP),can regulate BKCa activity and then elevate blood pressure.Methods Male Sprague-Dawley rats aged 6 weeks were randomized into high salt diet group(HS)and control group,fed with high salt diet(containing 5% NaCl)and standard rat chow(containing 0.4% NaCl)respectively for 16 weeks.Tail systolic blood pressure(SBP),body weight(BW)and 24-hour urinary output were tested every 4 weeks.Content of urinary Na+ was detected using flame spectrophotometrical method.At the end of 16 weeks,all the rats were killed,the mesenteric arteries were obtained,and single mesenteric smooth muscle cells were isolated at once.The resting membrane potential(Em),the total potassium currents and the currents after perfusion with TEA solution of the cells were all recorded by whole cell patch clamp.The transcriptions of BKCa channel α and β1 subunits in mesenteric arterial vascular smooth muscle cells(VSMC)of each group were calculated by real-time RT-PCR.Results There was no difference in SBP and BW at each stage between control group and HS group;the urinary Na+ level in HS animals was elevated significantly after 4 weeks.The negative values of Em in HS group VSMCs were reduced compared with those in the control group.Transcriptions of β1 subunit of BKCa channels were decreased in HS group,but α subunit transcriptions did not differ between the two groups.Whole cell potassium currents did not differ between HS and control groups,but BKCa currents of HS group VSMCs were lower than those of control group ones.Conclusion Even without elevating SBP,salt-loading can still modulate the expression and activity of BKCa channel in the mesenteric arterial VSMC and elevate vascular tone.

Tacrolimus Inhibits Vasoconstriction by Increasing Ca^(2+) Sparks in Rat Aorta

The present study attempted to test a novel hypothesis that Ca^2＋ sparks play an important role in arterial relaxation induced by tacrolimus. Recorded with confocal laser scanning microscopy, tacrolimus（10 μmol/L） increased the frequency of Ca^2＋ sparks, which could be reversed by ryanodine（10 μmol/L）. Electrophysiological experiments revealed that tacrolimus（10 μmol/L） increased the large-conductance Ca^2＋-activated K＋ currents（BKCa） in rat aortic vascular smooth muscle cells（AVSMCs）, which could be blocked by ryanodine（10 μmol/L）. Furthermore, tacrolimus（10 and 50 μmol/L） reduced the contractile force induced by norepinephrine（NE） or KCl in aortic vascular smooth muscle in a concentration-dependent manner, which could be also significantly attenuated by iberiotoxin（100 nmol/L） and ryanodine（10 μmol/L） respectively. In conclusion, tacrolimus could indirectly activate BKCa currents by increasing Ca^2＋ sparks released from ryanodine receptors, which inhibited the NE- or KCl-induced contraction in rat aorta.

The Involvement of Ca^2＋ Signal Pathways in Distal Colonic Myocytes in a Rat Model of Dextran Sulfate Sodium-induced Colitis

Background： Disrupted Ca2＋ homeostasis contributes to the development of colonic dysmotility in ulcerative colitis （UC）, but the underlying mechanisms are unknown. This study aimed to examine the alteration of colonic smooth muscle （SM） Ca2＋ signaling and Ca2＋ handling proteins in a rat model of dextran sulfate sodium （DSS）-induced UC. Methods： Male Sprague-Dawley rats were randomly divided into control （n = 18） and DSS （n = 17） groups. Acute colitis was induced by 5% DSS in the drinking water for 7 days. Contractility of colonic SM strips （controls, n = 8 and DSS, n = 7） was measured in an organ bath. Cytosolic resting Ca2＋ levels （n = 3 in each group） and Ca2＋ transients （n = 3 in each group） were measured in single colonic SM cells. Ca2＋ handling protein expression was determined by Western blotting （n = 4 in each group）. Differences between control and DSS groups were analyzed by a two-sample independent t-test. Results： Average tension and amplitude of spontaneous contractions of colonic muscle strips were significantly enhanced in DSS-treated rats compared with controls （1.25 ± 0.08 g vs. 0.96 - 0.05 g, P = 0.007; and 2.67 - 0.62 g vs. 0.52 ±0.10 g, P= 0.013）. Average tensions of carbachol-evoked contractions were much weaker in the DSS group （1.08 ±0.10 g vs. 1.80 ±0.19 g, P = 0.006）. Spontaneous Ca2＋ transients were observed in more SM cells from DSS-treated rats （15/30 cells） than from controls （5/36 cells）. Peak caffeine-induced intracellular Ca2＋ release was lower in SM cells of DSS-treated rats than controls （0.413 ±0.046 vs. 0.548 ±0.041, P = 0.033）. Finally, several Ca2＋ handling proteins in colonic SM were altered by DSS treatment, including sarcoplasmic reticulum calcium-transporting ATPase 2a downregulation and phospholamban and inositol 1,4,5-trisphosphate receptor 1 upregulation. Conclusions： Impaired intracellular Ca2＋ signaling of colonic SM, caused by alteration of Ca2＋ handing proteins, contribute to colonic dysmotility in DSS-induced UC.