Extracellular carbonic anhydrase of skeletal muscle associated with the sarcolemma

Abstract

We report here 1) the synthesis and properties of a new macromolecular carbonic anhydrase inhibitor, Prontosil-dextran, 2) its application to determine the localization of a previously described extracellular carbonic anhydrase in skeletal muscle, and 3) the application of a recently published histochemical technique using dansylsulfonamide to the same problem. Stable macromolecular inhibitors of molecular weights of 5,000, 100,000 and 1,000,000 were produced by covalently coupling the sulfonamide Prontosil to dextrans. Their inhibition constants towards bovine carbonic anhydrase II are 1-2 X 10(-7) M. The Prontosil-dextrans, PD 5,000, PD 100,000, and PD 1,000,000, were used in studies of the washout of H14CO3-) from the perfused rabbit hindlimb. This washout is slow due to the presence of an extracellular carbonic anhydrase and can be markedly accelerated by PD 5,000 but not by PD 100,000 and PD 1,000,000. Since PD 5,000 is accessible to the entire extracellular space and PD 100,000 and PD 1,000,000 are confined to the intravascular space, we conclude that the extracellular carbonic anhydrase of skeletal muscle is located in the interstitium. The histochemical studies show a strong staining of the sarcolemma of the muscle fibers with high oxidative capacity. It appears likely, therefore, that the extracellular carbonic anhydrase of skeletal muscle is associated with muscle plasma membranes with its active site directed toward the interstitial space.

Carbonic anhydrase in the sarcoplasmic reticulum of rabbit skeletal muscle.

“…This is in agreement with observations made with the histochemical technique of Hansson (L6nnerholm, 1980;Riley, Ellis & Bain, 1982). It is also in line with physiological studies indicating that the apparent volume of distribution of H14CO3-in the blood-free perfused rabbit hind limb is strongly reduced by a macromolecular carbonic anhydrase inhibitor that has access to the interstitial space but not by an inhibitor that is confined to the vascular bed (Gros, Geers, Dermietzel, Siffert, Gartner & Leibstein, 1984a, b;Geers, Gros & Gartner, 1985). It is not clear to what degree the carbonic anhydrase activity of the present s.r.…”

Mass spectrometric determination of HCO3− permeability and carbonic anhydrase activity in intact guinea‐pig colon epithelium

“…PMSF, DTT, ethoxzolamide (EthA) were purchased from Sigma; benzolamide (BA) was from Lederle (Pearl River, NJ, USA); and quaternary ammonium sulphonamide (QAS) was synthesized according to Wistrand (1984). Prontosil-Dextran 5000 (molecular weight, 5000) was prepared as described by Geers, Gros & G artner (1985).…”

“…It was therefore necessary to use extracellular carbonic anhydrase inhibitors to inhibit all extracellular carbonic anhydrase activity without affecting the intracellular enzyme. For this purpose we used Prontosil-Dextran 5000 (Geers, Gros & G artner, 1985) at a final concentration of 1 ² 10¦AE Ò, the relatively hydrophilic and membrane-impermeable benzolamide (BA) at a concentration of 1 ² 10¦AE Ò, or quaternary ammonium sulphonamide (QAS) (Wistrand, 1984) at a concentration of 1 ² 10¦Å Ò. At these concentrations all three inhibitors inhibited extracellular carbonic anhydrase activity completely (see Fig.…”

Acetazolamide, a Carbonic Anhydrase Inhibitor, Reverses Inflammation-Induced Thermal Hyperalgesia in Rats

“…It is proposed that CAs are involved in the extracellular control of pH in muscle (Wetzel et al, 2001). In the muscle, most of the subtypes of CAs (CA II, III, IV, and V) are found intracellularly, although there is at least one isoform, CA IV, that is found extracellularly on the sarcolemma (Geers et al, 1985;Decker et al, 1996;Geers and Gros, 2000). There is significant carbonic anhydrase activity in the extracellular space in the brain in rat hippocampal slice preparations (Chen and Chesler, 1992).…”