Title

The superficial buffer barrier hypothesis in venous smooth muscle

Date of Award

1994

Availability

Article

Degree Name

Doctor of Philosophy (Ph.D.)

Department

Molecular and Cellular Pharmacology

First Committee Member

David J. Adams, Committee Chair

Abstract

The interaction of Ca$\sp{2+}$ transport in the plasmalemma and the sarcoplasmic reticulum (SR) was investigated in smooth muscle of the rabbit inferior vena cava. SR depletion prior to stimulation of Ca$\sp{2+}$ influx induced a delay of 30-70 seconds between the intracellular Ca$\sp{2+}$ signal and force development. This delay was abolished by the application of caffeine. These data support the "Superficial Buffer Barrier" hypothesis, which holds that SR functions as a buffer barrier against Ca$\sp{2+}$ entry into the central cytoplasm.Thapsigargin completely blocked refilling of the SR regardless of Ca$\sp{2+}$ entry pathways, indicating that the SR Ca$\sp{2+}$-ATPase is essential for this process.Caffeine, ryanodine and thapsigargin, which inhibit SR Ca$\sp{2+}$ accumulation, increased the steady state intracellular Ca$\sp{2+}$ concentration ((Ca$\sp{2+}\rbrack\sb{\rm i}$). Measurements of Mn$\sp{2+}$ induced quench of the intracellular fura-2 signal during pharmacological manipulation of the SR content showed that these three agents did not stimulate divalent cation entry. On the other hand, stimulation with noradrenaline caused a marked increase in Mn$\sp{2+}$ influx, which was blocked by 2 mM Ni$\sp{2+}$. Mn$\sp{2+}$ entry stimulated by high K$\sp+$ solution was blocked by 1 $\mu$M diltiazem.After first raising (Ca$\sp{2+}\rbrack\sb{\rm i}$ with a high K$\sp+$, high Ca$\sp{2+}$ solution, subsequent replacement with a Ca$\sp{2+}$-free external solution will result in a decline in (Ca$\sp{2+}\rbrack\sb{\rm i}$. Comparison between the (Ca$\sp{2+}\rbrack\sb{\rm i}$ decays and their converted "rate-concentration" curves under different conditions shows that prior inhibition of SR Ca$\sp{2+}$ accumulation by caffeine or thapsigargin actually slows the rate of decline in (Ca$\sp{2+}\rbrack\sb{\rm i}$ which mostly reflects the process of Ca$\sp{2+}$ extrusion from the cells. There were no significant additive effects between the inhibition of Na$\sp+$/Ca$\sp{2+}$ exchanger and that of SR Ca$\sp{2+}$ accumulation. It is concluded that SR dysfunction has an inhibitory effect on Ca$\sp{2+}$ extrusion which results in an increase in the steady state (Ca$\sp{2+}\rbrack\sb{\rm i}$. The above results support the component of the "Superficial Buffer Barrier" hypothesis which proposes that ER Ca$\sp{2+}$ is discharged to the extracellular space via vectorial Ca$\sp{2+}$ release from the SR lumen.

Keywords

Health Sciences, Pharmacology; Biology, Animal Physiology

Link to Full Text

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