Awahan Rahman

Junctional and nonjunctional effects of heptanol and glycyrrhetinic acid derivates in rat mesenteric small arteries

Heptanol, 18α‐glycyrrhetinic acid (18αGA) and 18β‐glycyrrhetinic acid (18βGA) are known blockers of gap junctions, and are often used in vascular studies. However, actions unrelated to gap junction block have been repeatedly suggested in the literature for these compounds. We report here the findings from a comprehensive study of these compounds in the arterial wall. Rat isolated mesenteric small arteries were studied with respect to isometric tension (myography), [Ca2+]i (Ca2+‐sensitive dyes), membrane potential and – as a measure of intercellular coupling – input resistance (sharp intracellular glass electrodes). Also, membrane currents (patch‐clamp) were measured in isolated smooth muscle cells (SMCs). Confocal imaging was used for visualisation of [Ca2+]i events in single SMCs in the arterial wall. Heptanol (150 μM) activated potassium currents, hyperpolarised the membrane, inhibited the Ca2+ current, and reduced [Ca2+]i and tension, but had little effect on input resistance. Only at concentrations above 200 μM did heptanol elevate input resistance, desynchronise SMCs and abolish vasomotion. 18βGA (30 μM) not only increased input resistance and desynchronised SMCs but also had nonjunctional effects on membrane currents. 18αGA (100 μM) had no significant effects on tension, [Ca2+]i, total membrane current and synchronisation in vascular smooth muscle. We conclude that in mesenteric small arteries, heptanol and 18βGA have important nonjunctional effects at concentrations where they have little or no effect on intercellular communication. Thus, the effects of heptanol and 18βGA on vascular function cannot be interpreted as being caused only by effects on gap junctions. 18αGA apparently does not block communication between SMCs in these arteries, although an effect on myoendothelial gap junctions cannot be excluded.

Interaction Between Na+/K+-Pump and Na+/Ca2+-Exchanger Modulates Intercellular Communication

Ouabain, a specific inhibitor of the Na+/K+-pump, has previously been shown to interfere with intercellular communication. Here we test the hypothesis that the communication between vascular smooth muscle cells is regulated through an interaction between the Na+/K+-pump and the Na+/Ca2+-exchanger leading to an increase in the intracellular calcium concentration ([Ca2+]i) in discrete areas near the plasma membrane. [Ca2+]i in smooth muscle cells was imaged in cultured rat aortic smooth muscle cell pairs (A7r5) and in rat mesenteric small artery segments simultaneously with force. In A7r5 coupling between cells was estimated by measuring membrane capacitance. Smooth muscle cells were uncoupled when the Na+/K+-pump was inhibited either by a low concentration of ouabain, which also caused a localized increase of [Ca2+]i near the membrane, or by ATP depletion. Reduction of Na+/K+-pump activity by removal of extracellular potassium ([K+]o) also uncoupled cells, but only after inhibition of KATP channels. Inhibition of the Na+/Ca2+-exchange activity by SEA0400 or by a reduction of the equilibrium potential (making it more negative) also uncoupled the cells. Depletion of intracellular Na+ and clamping of [Ca2+]i at low concentrations prevented the uncoupling. The experiments suggest that the Na+/K+-pump may affect gap junction conductivity via localized changes in [Ca2+]i through modulation of Na+/Ca2+-exchanger activity.

Chronic cystamine treatment inhibits small artery remodelling in rats.

Background/Aims: We investigated whether the tissue transglutaminase inhibitor cystamine is able to inhibit remodelling of small arteries in vivo, a possibility suggested by recent in vitro experiments. Methods: Using osmotic minipumps, phenylephrine, cystamine and/or amlodipine were infused for 1–2 weeks into 9-week-old Wistar rats. Small arteries were then removed for pressure myograph investigation. Results: Phenylephrine infusion caused inward remodelling of the small arteries compared to vehicle infusion. The remodelling was abolished by concomitant infusion with cystamine; blood pressure was unaffected. Second, we investigated whether cystamine was able to inhibit outward remodelling. Rats were first infused with phenylephrine for 1 week, and some were infused for a further week with amlodipine with or without cystamine. Amlodipine caused 24% outward remodelling compared to vessels from rats at completion of the phenylephrine infusion. The outward remodelling was attenuated 86% by concomitant cystamine infusion. A series of in vitro experiments supported the inhibitory action of cystamine on tissue transglutaminase. Conclusion: The ability of cystamine to inhibit inward remodelling independent of blood pressure is consistent with a role of tissue transgluaminase in this process. It remains to be determined if the ability of cystamine to inhibit outward remodelling also involves inhibition of tissue transglutaminase.

Effects of cGMP on Coordination of Vascular Smooth Muscle Cells of Rat Mesenteric Small Arteries

Objective: We tested the hypothesis that cGMP can induce a state of only partial coordination of vascular smooth muscle cells (VSMC). Methods: This was done by studying the concentration-dependent effect of 8Br-cGMP on isometric and isobaric force development of noradrenaline-activated segments of rat mesenteric small arteries in which the endothelium was removed. We further measured the concentration-dependent effect of 8Br-cGMP on VSMC membrane potential, spatially resolved [Ca2+]i and VSMC membrane conductance. Results: With 300 µM 8Br-cGMP, coordinated [Ca2+]i activity and vasomotion were seen as previously reported. At 10–30 µM 8Br-cGMP, beating isometric tension oscillations were seen. Isobaric recordings revealed oscillations with different frequencies in different parts of the arteries. At these (10–30 µM) 8Br-cGMP concentrations, membrane potential oscillations did not always concur with isometric tension oscillations, and [Ca2+]i oscillations were only synchronized locally within groups of cells. 8Br-cGMP concentration-dependently decreased the frequency of vasomotion and, in unsynchronized hyperpolarized VSMC, the frequency of [Ca2+]i waves. Conclusion: Our results demonstrated that cGMP can cause a partial coordination of the VSMC in the vascular wall (and at high concentrations near complete coordination). Furthermore, the cGMP concentration-dependent decrease of Ca2+ wave frequency and of vasomotion frequency suggests that cGMP modifies oscillatory Ca2+ release from the sarcoplasmic reticulum and supports the suggestion that this oscillatory release paces vasomotion.

Thanks to the Reviewers

CHRISTOPHER ADAMSON MICHAEL P. ALLEN WILLIAM BAILEY STEVE BALKIN RICHARD E. BALL JOYCE BARAKETT ROBERT A. BEAUREGARD E. M. BECK HOWARD S. BECKER LAWRENCE BENNETT BENNETT BERGER LEONARD BERKEY MICHAEL BETZ EGON BITTNER LEONARD BLUMBERG PHILIP BLUMSTEIN ROBERT BOGDEN EDNA BONACICH PEG BORTNER VERN BULLOUGH LEONARD CAIN KITTY CALAVITA JOHN CARSLEY JOAN CASSELL PAUL CHALFANT RANDALL COLLINS MARK COLVIN JOHN A. CONLEY ELAINE CUMMING ARLENE KAPLAN DANIELS JON DARLING JOAN DEBARDELEBEN CHARLES DERBER STEVEN DEUTSCH IRWIN DEUTSCHER PAUL DIMAGGIO JOAN EAKIN WILLIAM EATON SUSAN ECKSTEIN SHELDON EKLAND-OLSON ROBERT EMERSON PAULA S. ENGLAND DAVID M. ERMANN DARYL EVANS WILLIAM FALK SAMUEL FARBER ROB FAULKNER JOE FEAGIN ROSLYN FELDBERG SANDY FELDHEIM MARK FISHMAN NANCY FRANK ELIOT FREIDSON PETER FREITAG EDGAR Z. FRIEDENBERG WILLIAM FRIEDLAND SAMUEL FRIEDMAN JOHN GALLIHER DAVID GARTMAN GILBERT GEIS JAMES GESCHWENDER DON C. GIBBONS PEGGY GIORDANO DANIEL GLASER FRED GOLDNER ROBERT A. GORDON WALTER GOVE SUSAN GRAY DAVID GREENBERG ALLEN D. GRIMSHAW EDWARD GROSS BRUCE HACKETT JOHN HAGAN RICHARD F. HAMILTON SUE KIEFER HAMMERSMITH SHARON HARLAN CLAYTON HARTJEN JAMES HENSLIN JOHN P. HEWITT BARBARA HEYL SALLY T. HILLSMAN LYNDA LYTLE HOLMSTROM RANDY HODSON ALLEN HORWITZ JOAN HUBER DREW HUMPHRIES ALLEN W. IMERSHEIN JAMES INCIARDI DAVID JACOBS GARY JENSEN CAROLE JOFFE JOHN JOHNSON PAUL JOSEPH RACHEL KAHN-HUT DEBRA KALMUSS JACK KATZ RONALD KESSLER JOSEPH A. KOTARBA RONALD KRAMER NANCY KUTNER BARBARA R. LASLETT PAT LAUDERDALE RONALD LAWSON JEFFREY LEITER RHONDA F. LEVINE CYRIL LEVITT ELLIOT LIEBOW CLARENCE LO ULI LOCHER MARGARET LOCK HELENA Z. LOPATA DAVID E. LOPEZ DAVID LUCKENBILL PETER LYMAN LARRY LYON STEVE LONGSTAFF CHARLES McCAGHY JOHN D. McCARTHY SCOTT G. McNALL LINDA C. MAJKA GERALD MARKLE JOHN MARKOFF CORA MARRETT TONY MASI DOUGLAS MAYNARD ROBERT MEIER JANET E. MICKISH TERANCE MIETHE S.M. MILLER BETH MINTZ MERRY ANN MORASH DAVID L. MORGAN CAROL KIAPERMAN MORROW CHARLES C. MOSCOS JR MARTHA MYERS JOANE NAGEL DOROTHY NELKIN MARGARET K. NELSON LINDA B. NILSON STEPHEN NORLAND MELVIN OLIVER MARVIN OLSEN SUSAN OLZAK ANN L. PAGE TOBY PARCEL DOROTHY PAWLUCH HAROLD E. PEPINSKY CHARLES PERROW ROBERT PERRUCCI SUZANNE PETERS KAREN J. PETERSON MICHAEL PETRUNIK MARK PEYROT GERALD PLATT ALPHONSO PINKNEY HENRY PONTELL ROBERT C. PRUS RICHARD QUINNEY NICOLE FISCHER RAFTER PRUDENCE RAINS DONNA RANDALL STEVEN RANDALL JOSEPH RANKIN RICHARD RATCLIFF PAMELA RICHARDS JAMES RICHARDSON RAY RIST JACK ROACH JANET ROACH JIM ROBBINS E. BURKE ROCHFORD JOSEPH W. ROGERS ROBIN ROOM RACHEL ROSENFELD BARBARA KATZ ROTHMAN LILLIAN RUBIN JOSEPHINE A. RUGGIERO WILLIAM A. RUSHING SHERYL RUZEK SASKIA SASSEN-KOOB SANDRA P. SCHOENBERG WILLIAM SHAFFIR NANCY S. SHAW CLIFFORD D. SHEARING JAMES F. SHORT JR. ROBERTA SIMMONS DOUGLAS SMITH MICHAEL SMITH DAVID A. SNOW LEE SODERSTROM JACK W. SPENCER STEVEN SPITZER DARRELL J. STEFENMEIER RONALD TAYLOR JIM THOMAS ROBERT THOMAS GARY TIEDEMAN KATHLEEN TIERNEY RONALD TROYER JAMES D. UNNEVER BERT USEEM PAULINE VAILLANCOURT ARTHUR J. VIDICH DIANA VAUGHAN EDWARD J. WALSH VIVIENNE WALTERS CATHERINE WATSON J. ALLEN WHITT SIDNEY WILLHELM JUDITH G. WITTNER PETER YIN MAYER N. ZALD DAVID ZARET