Lactate metabolism in the perfused rat hindlimb

Abstract

A preparation of isolated rat hindleg was perfused with a medium consisting of bicarbonate buffer containing Ficoll and fluorocarbon, containing glucose and/or lactate. The leg was electrically prestimulated to deplete partially muscle glycogen. The glucose was labelled uniformly with 14C and with 3H in positions 2, 5 or 6, and lactate uniformly with 14C and with 3H in positions 2 or 3. Glucose carbon was predominantly recovered in glycogen, and to a lesser extent in lactate. The 3H/14C ration in glycogen from [5-3H,U-14C]- and [6-3H,U-14C]-glucose was the same as in glucose. Nearly all the utilized 3H from [2-3H]glucose was recovered as water. Insulin increased glucose uptake and glycogen synthesis 3-fold. When the muscle was perfused with a medium containing 10 mM-glucose and 2 mM-lactate, there was little change in lactate concentration. 14C from lactate was incorporated into glycogen. There was a marked exponential decrease in lactate specific radioactivity, much greater with [3H]- than with [14C]-lactate. The 'apparent turnover' of [U-14C]lactate was 0.28 mumol/min per g of muscle, and those of [2-3H]- and [3-3H]-lactate were both about 0.7 mumol/min per g. With 10 mM-lactate as sole substrate, there was a net uptake of lactate, at a rate of about 0.15 mumol/min per g, and the apparent turnover of [U-14C]lactate was 0.3 mumol/min per g. The apparent turnover of [3H]lactate was 3-5 times greater. When glycogen synthesis was low (no prestimulation, no insulin), the incorporation of lactate carbon into glycogen exceeded that from glucose, but at high rates of glycogen deposition the incorporation of lactate carbon was much less than that of glucose. Lactate incorporation into glycogen was similar in fast-twitch white and fast-twitch red muscle, but was very low in slow-twitch red fibres. We find that (a) pyruvate in muscle is incorporated into glycogen without randomization of carbon, and synthesis is not inhibited by mercaptopicolinate or cycloserine; (b) there is extensive lactate turnover in the absence of net lactate uptake, and there is a large dilution of 14C-labelled lactate from endogenous supply; (c) there is extensive detritiation of [2-3H]- and [3-3H]-lactate in excess of 14C utilization.

Dichloroacetate inhibits glycolysis and augments insulin-stimulated glycogen synthesis in rat muscle.

“…Because glucose oxidation was increased by only 0.5-1.5 ,umol/g per h, > 80% of the decrease in lactate release caused by DCA resulted from inhibition of net glycolysis. To study the mechanism by which DCA reduced glucose uptake at supraphysiologic levels of insulin, glucose transport and phosphorylation were measured in muscles of fasted rats using 2-deoxy-[U-'4C]glucose and [2-3H]glucose (9,11,12). With 10 mU/ml insulin, rates of glucose transport and phosphorylation were the same (Table I); when DCA was added, 2-deoxy-[U-'4C]glucose transport was unchanged, but glucose phosphorylation (the rate of 3H20 formed) was decreased 17% after 30 min and 12% after 2 h of incubation.…”

“…Studies by McLane and Holloszy (34) and Shiota et al (12) have demonstrated that skeletal muscle from fed rats can incorporate significant amounts of lactate into glycogen when the extracellular lactate concentration is high. We confirmed this and found that this pathway was depressed by fasting (Fig.…”

“…2). Calculation ofglycogen synthesis using the extracellular specific radioactivity of lactate can underestimate the rate of glycogen synthesis from lactate (12,34). Measurement of the intracellular specific radioactivity of pyruvate would be preferable, but to do this accurately in muscles of the size we studied is not possible (35).…”

“…Glucose [2-3H]glucose (9). The formation of 3H20 was measured because it is an index ofthe rate ofglucose phosphorylation ( 11) and corresponds closely to glucose uptake by perfused muscle (12 Glucose uptake was calculated as [5-3H]glucose incorporated into glycogen plus 3H20 formed during glycolysis (9). Net glycolysis was calculated as the difference between the rate of 3H20 formed and substrate recycling (15).…”

NIRS Measurement of O2 Dynamics in Contracting Blood and Buffer Perfused Hindlimb Muscle

“…1 shows the system. Surgery was modified from those in previous reports [11][12][13]. The inflow catheter was placed in the descending aorta with its tip 4-5 mm proximal to the aorta bifurcation.…”