CK inhibition accelerates transcytosolic energy signaling during rapid workload steps in isolated rabbit hearts

Abstract

The effect of graded creatine kinase (CK) inhibition on the response time of mitochondrial O2 consumption to dynamic workload jumps ( t mito) was studied in isolated rabbit hearts. Tyrode-perfused hearts ( n = 7/group) were exposed to 15 min of 0, 0.1, 0.2, or 0.4 mM iodoacetamide (IA) (CK activity = 100, 14, 6, and 3%, respectively). Pretreatment t mito was similar across groups at 6.5 ± 0.5 s (mean ± SE). The increase observed over time in control hearts (33 ± 8%) was progressively reversed to 16 ± 6, −20 ± 6 ( P< 0.01 vs. control), and −46 ± 6 ( P < 0.01 vs. control) % in the 0.1, 0.2 and 0.4 mM IA groups, respectively. The faster response times occurred without reductions in mitochondrial oxidative capacity (assessed in vitro) or myocardial O2 consumption of the whole heart during workload steps. Isovolumic contractile function assessed as rate-pressure product (RPP) and contractile reserve (increase in RPP during heart rate steps) were significantly reduced by IA. We conclude that CK in the myofibrils and/or cytosol does not speed up transfer of the energy-related signal to the mitochondria but rather acts as an energetic buffer, effectively slowing the stimulus between myofibrils/ion pumps and oxidative phosphorylation. This argues against the existence of an obligatory creatine phosphate energy shuttle, because CK is effectively bypassed.

Glycolytic buffering affects cardiac bioenergetic signaling and contractile reserve similar to creatine kinase

“…As shown, there was no difference in any parameter between groups' pretreatment, but significant systolic and diastolic impairment after inhibition of glycolysis. Systolic left ventricular pressure was significantly decreased in the IAA group, whereas end-diastolic pressure was elevated in both the IA-and IAA-treated hearts, findings consistent with our and others previous use of these compounds (10,15,32,34) and potentially reflecting an increase in the free cytosolic [ADP]. Contractile function stabilized after the washout period and no significant deterioration was observed during the second series of heart rate steps.…”

“…In addition, the change in preto posttreatment t mito observed in IA-treated hearts was significantly larger than that in the IAA group. Figure 2 combines the posttreatment values of t mito in the current study to those found in our earlier study (10), in which glucose alone was used as substrate and IA was infused at a range of concentrations (0.1, 0.2, and 0.4 mM; only the latter is shown for clarity). Figure 2 illustrates the accelerating effect of pyruvate addition alone to the buffer on t mito in control hearts.…”

“…t transport , which characterizes oxygen transport, was not affected by the IA or IAA infusion (see Table 3). We have previously shown in mitochondria isolated from 0.4 mM IA-treated hearts that state 3 and 4 V O 2 stimulated with ADP is unaffected, suggesting no damage to oxidative phosphorylation in these hearts (10). State 3 respiration in isolated mitochondria was not affected by addition of up to 1.5 mM IA or up to 1 mM IAA, appreciably higher concentrations than used during perfusion.…”

“…This inhibition was achieved by the differential infusion of the sulfhydryl-enzyme blockers iodoacetamide (IA) and iodoacetic acid (IAA), used previously by us (10) and by others (15,23,(32)(33)(34) to preferentially reduce CK and GAPDH activity, respectively. On the basis of our preliminary findings (36), we predicted that the removal of ATP and ADP buffering capacity of glycolysis alone or in combination with CK inhibition would speed up signaling to oxidative phosphorylation at the expense of a compromised ability to perform acute increases in myocardial workload.…”

“…IAA at 0.3 mM was more effective against GAPDH (2% of control remaining) versus CK (80% of control). Because of the reserve in the CK system, CK activity alone has to be reduced to Ͻ15% of control to cause any changes in contractile function and energy signaling speeds as shown previously (10,34). IAAtreated hearts are therefore glycolytically blocked but CK operable.…”