METABOLIC STUDIES OF CREATINE ANALOGUES; ATP-SPARING DURING ISCHEMIA IN HEART AND SKELETAL MUSCLE OF CHICKS FED CYCLOCREATINE
TURNER, DAVID M.
Doctor of Philosophy
Several embryonic tissues were utilized in the study of creatine analogues. Creatine analogues were taken up and phosphorylated by incubated chick embryo muscle, skin, and heart, in addition to the previously studied chick embryo brain. Accumulated phosphagen was utilized during insult by ischemia or by the addition of various inhibitory compounds to media containing chick embryonic tissues. Resynthesis of the phosphagen, upon cessation of insult and return of the tissues to the incubation media, usually occurred. The enzyme arginine:glycine amidinotransferase was partially purified for the first time from liver. Through the use of several chromatographic techniques a 160-fold purification of the enzyme was achieved from chick liver. Hearts of chicks fed 1-carboxymethyl-2-iminoimidazolidine (cyclocreatine) accumulated 15 (mu)mol/g wet weight of cyclocreatine-P and had glycogen levels elevated three-fold relative to controls. During total ischemia in vitro, cyclocreatine-P was utilized by heart for resynthesis of ATP, glycolysis was greatly prolonged and depletion of both ATP and total adenylate pools was delayed two to five-fold relative to controls. Feeding of a second analogue, 1-carboxyethyl-2-iminoimidazolidine (homocyclocreatine) resulted in the accumulation of homocyclocreatine-P in the heart and a tripling of the glycogen levels. During ischemia, homocyclocreatine-P was only slightly utilized, glycolysis was prolonged, and most importantly, no delay in depletion of ATP levels was observed relative to controls. Feeding of creatine resulted in a doubling of creatine levels in the heart but no delay of ATP depletion during ischemia relative to controls. Maximal conservation of ATP and total adenylates in the heart during ischemia required, in addition to adequate glycogen reserves, a phosphagen which is both kinetically and thermodynamically competent to buffer ATP/ADP during the later stages of ischemia; that is, cyclocreatine-P. Breast muscle of chicks fed cyclocreatine demonstrated a decreased rate of depletion of glycogen upon exposure to the (beta)-adrenergic-agonist, isoproterenol, relative to controls. In addition, during ischemia following exposure to isoproterenol, breast muscle from cyclocreatine-fed chicks was protected against depletion of ATP, with ATP levels remaining 6-fold elevated after 2 hours of ischemia relative to controls. A correlation was demonstrated between total cyclocreatine accumulated and sparing of ATP during ischemia.