Pyruvate represents an important branch point in carbohydrate metabolism and its fate depends (in part) upon the oxidation state of the cell. In cardiac muscle, pyruvate
produced by glycolysis may be converted to acetyl-CoA, lactate or alanine, though the latter normally represents a minor route of metabolism.
The oxidative decarboxylation of pyruvate to acetyl-CoA and CO2 is catalysed by the multienzyme pyruvate dehydrogenase (PDH) complex. The mammalian PDH complex contains multiple copies of; pyruvate dehydrogenase (El), dihydrolipoamide acetyltransferase (E2) and dihydrolipoamide dehydrogenase (E3) which together catalyse the decarboxylation reactions; two regulatory enzymes (PDH kinase and PDH phosphatase); and protein X which links E3 to the complex (Patel & Roche, 1990). The activity of the PDH complex is primarily regulated by reversible protein phosphorylation. Phosphorylation of E l by the intrinsic PDH kinase results in inactivation, and there are no know activators of the phosphorylated complex (PDHy) (Randle et al, 1994). Dephosphorylation and reactivation is catalysed by the PDH
phosphatase. Phosphorylation can occur at multiple sites on E l and this retards reactivation of the complex by PDH phosphatase in vitro (Sugden & Holness, 1994).
The total amount of PDH complex is constant under most physiological conditions, but the functional activity varies widely depending on the relative proportion of the complex in the active, dephosphorylated form (PDHa). At rest, approximately 20% of the complex is in the active form, but this may increase to 90% when glycolysis is stimulated, for example during exercise (Opie, 1991). PDH kinase activity is stimulated by increasing mitochondrial ratios of acetyl-CoA/CoASH, NADH/NAD^ and ATP/ADP. The activating effects of acetyl-CoA and NADH involve reduction of the lipoyl moiety o f the complex, and are antagonised by Co ASH and NAD^ respectively (Randle et al,
1994). PDH kinase is also inhibited by pyruvate which therefore acts as a feed-forward activator of the PDH complex. The cardiac PDH is highly sensitive to activation by pyruvate, which may facilitate the use of exogenous lactate, explaining why this is a
preferred fuel of the heart (Sugden & Holness, 1994; Hue et al, 1995). PDH
phosphatase is dependent upon and is activated by physiological concentrations of Ca^^ (O.l-lOpM) (Randle et al, 1994). Perfiision of isolated hearts with increasing Ca^^
concentrations has been shown to increase the amount of PDH in the active, dephosphorylated form (McCormack et al, 1990). PDH phosphatase may be inhibited by
NADH (Randle et al, 1994).
Perfusion of rat hearts with adrenaline increases the amount o f PDHa probably by increasing cellular Ca^^ levels, which in turn activates PDH phosphatase. The elevated cytoplasmic (and thus mitochondrial) Ca^^ concentration associated with muscle contraction may be a major factor responsible for the increase in PDHa induced by increased cardiac workload (McCormack et al, 1990; Randle et al, 1994). Cardiac work
may also decrease the mitochondrial acetyl-Co A/Co ASH and NADH/NAD^ ratios, thereby relieving inhibition of PDH by PDH kinase. Activation of PDH phosphatase by insulin as observed in adipose tissue has not been demonstrated in cardiac muscle (Randle et al, 1994), although insulin infusion increases the amount o f active cardiac
PDHa in vivo (Cooney et al, 1993).
Perfusion of rat hearts with fatty acids or ketone bodies decreases the proportion of PDH in the active form by up to 70% (Randle et al, 1994), demonstrating the
importance o f PDH in the 'glucose-fatty acid cycle'. Such effects are probably mediated via the activation of PDH kinase by the rise in mitochondrial acetyl-CoA/CoASH and NADH/NAD^ ratios brought about by oxidation of these lipid fuels.
1.3.1.6 Glycogen
Glycogen present in cytoplasmic storage granules is also a potential source of glucose for myocardial energy generation. The regulation of cardiac muscle glycogen
metabolism and its physiological function are not well understood. Glycogen synthesis and degradation occur by separate pathways which in skeletal muscle are regulated reciprocally as outlined below. Incorporation of glucose-6-phosphate into glycogen involves three steps the third of which, catalysed by glycogen synthase (GS), is regulatory. GS activity is controlled by interconversion between an 'active', dephosphorylated form (GSa) and an 'inactive', phosphorylated form (GSb). Rat heart GS
has recently been purified to apparent homogeneity (~87 kDa by SDS-PAGE) and can be deactivated via phosphorylation by a variety of kinases in vitro, including GS kinase 3
and PKA (Grekinis et al, 1995). Dephosphorylation and reactivation may be brought
about by the action of several phosphatases including protein phosphatase-1. Glycogen phosphorylase catalyses glycogen breakdown and is also regulated by reversible phosphorylation. Phosphorylation activates glycogen phosphorylase and is catalysed by a specific phosphorylase kinase, which itself is activated through phosphorylation by PKA. Thus a rise in cellular cAMP levels, caused by p-adrenergic stimulation, initiates a cascade of events leading to activation of glycogen phosphorylase and inhibition of GS. In addition, protein phosphatase inhibitor-1 (PPI-1) present in cardiomyocytes has been shown to be hormonally regulated through activation by PKA (Gupta et al, 1996). Thus,
cAMP also down-regulates glycogen synthesis via PPI-1, which inhibits protein phosphatase-1 whose actions would tend to restore GS activity.
Glycogen metabolism is also regulated by non-hormonal factors. Phosphorylated GSb is activated by glucose-6-phosphate but inhibited by ATP and Pj, whilst GSa is active in the absence of glucose-6-phosphate and only weakly inhibited by ATP and Pi. Inactive glycogen phosphorylase is stimulated by AMP.
Anoxia and increased cardiac workload stimulate mobilisation of glycogen and inhibit its synthesis (presumably via a fall in high energy phosphates and rise in AMP and P|).
Glycogen is rapidly resynthesised following depletion of endogenous stores by intense work or ischaemia, indicating that low glycogen levels promote glycogen synthesis (Opie, 1991). Endogenous glycogen may serve as a substrate reservoir to buffer rapid increases in energy demand. In isolated working rat hearts, glycogen appears to be preferentially oxidised (as opposed to being metabolised to lactate) and it has been suggested that this ensures efficient generation of ATP from a limited supply of endogenous substrate (Goodwin etal, 1996).