Phosphate uptake and PiT-1 protein expression in rat skeletal muscle

Abstract

Skeletal muscle fiber types differ in their contents of total phosphate, which includes inorganic phosphate (P i) and high-energy organic pools of ATP and phosphocreatine (PCr). At steady state, uptake of Pi into the cell must equal the rate of efflux, which is expected to be a function of intracellular P i concentration. We measured 32 P-labeled Pi uptake rates in different muscle fiber types to determine whether they are proportional to cellular Pi content. Pi uptake rates in isolated, perfused rat hindlimb muscles were linear over time and highest in soleus (2.42 Ϯ 0.17, and lowest in white gastrocnemius (0.49 Ϯ 0.06Reasonably similar rates were obtained in vivo. Pi uptake rates at plasma Pi concentrations of 0.3-1.7 mM confirm that the P i uptake process is nearly saturated at normal plasma Pi levels. Pi uptake rate correlated with cellular Pi content (r ϭ 0.99) but varied inversely with total phosphate content. Sodium-phosphate cotransporter (PiT-1) protein expression in soleus and red gastrocnemius were similar to each other and seven-to eightfold greater than PiT-1 expression in white gastrocnemius. That the PiT-1 expression pattern did not match the pattern of P i uptake across fiber types implies that other factors are involved in regulating Pi uptake in skeletal muscle. Furthermore, fractional turnover of the cellular P i pool (0.67, 0.57, and 0.33 h Ϫ1 in soleus, red gastrocnemius, and white gastrocnemius, respectively) varies among fiber types, indicating differential management of intracellular Pi, likely due to differences in resistance to Pi efflux from the fiber. inorganic phosphate; sodium-inorganic phosphate transporters; PiT-2; inorganic phosphate efflux IN EVERY CELL, phosphate is necessary for structural and metabolic needs. In cellular energetics, phosphate forms the highenergy bonds of ATP and phosphocreatine (PCr), and inorganic phosphate (P i ) is a substrate for reactions in glycolysis, tricarboxylic acid cycle, and mitochondrial F 0 F 1 ATPase. Because cellular P i is a determinant of the free energy of ATP hydrolysis, its concentration is tightly controlled. However, alterations in total phosphate content in skeletal muscle, e.g., expansion of the PCr and/or reduction of the ATP pools, suggest that either P i uptake or P i loss from the cell is also modulated.In skeletal muscle, phosphate is distributed among the P i , ATP, and PCr pools primarily via ATPases, mitochondrial oxidative phosphorylation, and the creatine kinase reaction (21). Maintenance of the total phosphate contained in these pools depends on the balance of uptake and efflux of phosphate across the sarcolemma. Although most cellular phosphate is contained in organic phosphates, these molecules cannot easily cross the plasma membrane; rather, phosphate is transported as P i . Because P i is transported against a concentration and electrical gradient, it must be imported via active transport. This is presumably accomplished by the type III sodiumphosphate (Na-P i ) cotransporter (15) and is driven by the electrochemical ...

1H NMRS of carnosine combined with 31P NMRS to better characterize skeletal muscle pH dysregulation in Duchenne muscular dystrophy

“…44 We also mention a possible role of Na + /P i cotransporters. 45 The increased pH values determined by 1 H NMRS, as found in this study in at least a proportion of patients with DMD, strongly support the hypothesis of a cytoplasmic pH dysregulation in leaky suffering myocytes. Although there is no splitting of the carnosine C2-H resonance, it is significantly broader in the case of elevated 1 H-derived pH, favoring a two-pool scenario: one with a neutral pH and one with an alkaline pH.…”

Phosphate uptake in rat skeletal muscle is reduced during isometric contractions

“…Before use, the perfusate was warmed to 37°C and pH was adjusted to 7.40. Standard P i concentration in Krebs-Henseleit is 1.1 mM, a concentration shown previously to effectively saturate the uptake process (1).…”

Regulation of mitochondrial respiration by inorganic phosphate; comparing permeabilized muscle fibers and isolated mitochondria prepared from type-1 and type-2 rat skeletal muscle

“…Intracellular concentration of P i has been reported to vary between 3.1 mM and 10.1 mM in cat biceps and soleus (Meyer et al 1985), 1.1 mM in rat EDL and 3.6 mM in rat soleus (Kushmerick et al 1992), 3.6, 2.3 and 1.5 mol/g in rat soleus, red gastrocnemius and white gastrocnemius (Abraham et al 2004) at rest. This yields a possible 10-20% increase in respiration when phosphate increases from the resting value to 10 mM and beyond.…”