Activation and measurement of plasma prorenin in the rat.
Prorenin determination in rat plasma has been problematic from the outset. Consequently, its existence is questioned by some and its quantity by others, making it difficult for knowledge to advance as to its function relative to the renin system. The present study examines major variables in the determination of rat plasma prorenin and renin, notably different prorenin activation protocols involving blood samples obtained under various conditions from animals under different anesthetics. We found that a trypsin activation step with 5 mg/mL plasma, 60 min at 23 degrees C, followed by a PRA step of 10 min at 37 degrees C, resulted in the highest prorenin estimates, up to approximately 400 ng.mL-1.h-1 in terms of angiotensin I, as compared with published values of 0-190, based on other protocols. These estimates were obtained despite considerable destruction of angiotensinogen (renin substrate) by trypsin. Cryoactivation of prorenin was much less effective than in human plasma but, when followed by trypsin, it facilitated greater activation than with trypsin alone. Comparable fresh and fresh-frozen plasmas had similar prorenin-renin values, but lower values were observed in plasmas that had been repeatedly frozen and thawed. Conscious rats and those anesthetized with Inactin or ether had higher renins and prorenins than those anesthetized with methoxyflurane or halothane. Rats with kidneys in place during blood collection had higher renins (but not prorenins) than those whose kidneys were clamped off, suggesting that last-minute renin release during blood collection had occurred. We conclude that (i) trypsin generates increased renin, or renin-like, activity in plasma, suggesting activation of a precursor; (ii) on this basis, high prorenin levels exist in normal rat plasma; (iii) renin and prorenin levels are variously influenced by different anesthetics and blood handling procedures; (iv) variation in prorenin levels suggests that it is a dynamic (functional?) component of the renin system; (v) prorenin measurements are heavily influenced by methodological variations during the trypsin step or the subsequent PRA step; (vi) using standardized methodology, the rat can serve as a model for investigating the function of prorenin in normotension and hypertension.
Ioannou
P
P
Department of Physiology, University of Toronto, Ont., Canada.
Loh
A Y
AY
Osmond
D H
DH
D000311:Adrenal Glands / Q000502:physiology; D000777:Anesthetics; D000804:Angiotensin II / Q000097:blood; D000818:Animals; D001800:Blood Specimen Collection / Q000379:methods; D004789:Enzyme Activation / Q000502:physiology; D004792:Enzyme Precursors / Q000097:blood* / Q000378:metabolism; D005615:Freezing; D006973:Hypertension / Q000201:enzymology*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D012083:Renin / Q000097:blood* / Q000378:metabolism; D014357:Trypsin / Q000494:pharmacology
D000777:Anesthetics; D004792:Enzyme Precursors; D000804:Angiotensin II; D014357:Trypsin; D012083:Renin