Isolation and purification of a vascular hyperreactivity factor from rabbit kidney cortex
Williams, Warren Jon-Allan
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https://hdl.handle.net/2142/22683
Description
Title
Isolation and purification of a vascular hyperreactivity factor from rabbit kidney cortex
Author(s)
Williams, Warren Jon-Allan
Issue Date
1990
Doctoral Committee Chair(s)
Zehr, John E.
Department of Study
Molecular and Integrative Physiology
Discipline
Molecular and Integrative Physiology
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Animal Physiology
Language
eng
Abstract
Altered vascular response to agonist has been a consistently observed phenomenon in experimental animal and human hypertension. Much evidence suggests that a circulating factor of renal origin is, in part, responsible for this vascular hyperreactivity. The focus of this study was to isolate, purify and identify a vascular hyperreactivity factor (VHRF) from rabbit kidney cortex. Further studies were performed to define the mechanism of action of the VHRF in muscle.
Renal cortical tissue was obtained from euthanized New Zealand White rabbits (Oryctolagus cunniculus). By application of a series of techniques (dialysis, gel and ion exchange chromatography, preparative electrofocusing, HPLC, FAB mass spectrometry, FT infrared spectrometry) an acid stable, low molecular weight (147 daltons) compound with a high (+) charge density was obtained. When injected (10 $\mu$g, i.v.) into assay rats, this compound amplified the pressor response to fixed dosages (5-10 $\mu$g) of norepinephrine (NE). Taken together, this compound exhibits nearly identical physical characteristics (acid stability, structure, charge and biological activity) to the naturally occurring polyamine spermidine (145.6 daltons). Moreover, in dose-response experiments, $\mu$g dosages of spermidine amplified the pressor response to a range (5-25 $\mu$g) of dosages of NE.
The mechanism of action of spermidine was examined in a series of experiments using frog semitendinosus muscle fibers. Single fibers were isolated, mechanically skinned, suspended between a micrometer and a photoelectric force transducer and immersed in a tissue bath. Fiber diameter was recorded at slack length and then a 10% stretch applied. Initially, fiber tension was induced by depolarization with 92 mM chloride. Tension development was found to be inhibited in a dose-dependent manner by $\mu$M concentrations of spermidine. However, when fiber tension was induced by 2.5 mM caffeine with $\mu$M concentrations of spermidine, tension development by caffeine was enhanced compared to control values. When fiber tension was induced incrementally by increasing the calcium concentration (pCa tension curve) the presence of spermidine did not displace the pCa tension curve. These experiments suggest that spermidine is acting on the ryanodine receptor to potentiate calcium release from the sarcoplasmic reticulum and does not act either on the dihydropyridine receptor or the contractible elements of the fiber.
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