TY - JOUR
T1 - A physiological measure of carbonic anhydrase in müller cells
AU - Newman, Eric A.
PY - 1994/8
Y1 - 1994/8
N2 - Carbonic anhydrase activity was characterized in freshly dissociated Müller cells of the salamander retina. Intracellular pH was monitored using ratio imaging of the indicator dye BCECF as extracellular Pco2 was varied. The extracellular solution was switched rapidly (141 ms rise time) from a HEPES buffered to a CO2‐HCO−3 buffered solution (both pH 7.4). Introduction of CO2‐HCO−3 produced a rapid cell acidification. Cell pH dropped from a steady‐state pH of 7.02 in HEPES solution to pH 6.81 in CO2‐HCO−3. Methazolamide, a carbonic anhydrase inhibitor, dramatically reduced the initial rate of acidification, demonstrating that the acidification was produced by the carbonic anhydrase‐catalyzed hydration of CO2. The initial rate of acidification, 52.6 pH units per min (0.88 pH units per s), was reduced ∼ 150‐fold to 0.36 pH units per min by 10−3 M methazolamide. Half‐maximal inhibition occurred at a methazolamide concentration of 5.6. 10−7 M. The carbonic anhydrase inhibitor acetazolamide (10−3 M) also greatly reduced the rate of cell acidification. The latency to the onset of carbonic anhydrase inhibition was 660 ms for methazolamide and 7.5 s for acetazolamide. The carbonic anhydrase inhibitor benzolamide (10−4 M, 4 min exposure), which is poorly membrane permeant, had little effect on the rate of cell acidification, indicating that the site of carbonic anhydrase action was intracellular. The activity of Müller cell carbonic anhydrase may help to buffer extracellular CO2 variations in the retina. © 1994 Wiley‐Liss, Inc.
AB - Carbonic anhydrase activity was characterized in freshly dissociated Müller cells of the salamander retina. Intracellular pH was monitored using ratio imaging of the indicator dye BCECF as extracellular Pco2 was varied. The extracellular solution was switched rapidly (141 ms rise time) from a HEPES buffered to a CO2‐HCO−3 buffered solution (both pH 7.4). Introduction of CO2‐HCO−3 produced a rapid cell acidification. Cell pH dropped from a steady‐state pH of 7.02 in HEPES solution to pH 6.81 in CO2‐HCO−3. Methazolamide, a carbonic anhydrase inhibitor, dramatically reduced the initial rate of acidification, demonstrating that the acidification was produced by the carbonic anhydrase‐catalyzed hydration of CO2. The initial rate of acidification, 52.6 pH units per min (0.88 pH units per s), was reduced ∼ 150‐fold to 0.36 pH units per min by 10−3 M methazolamide. Half‐maximal inhibition occurred at a methazolamide concentration of 5.6. 10−7 M. The carbonic anhydrase inhibitor acetazolamide (10−3 M) also greatly reduced the rate of cell acidification. The latency to the onset of carbonic anhydrase inhibition was 660 ms for methazolamide and 7.5 s for acetazolamide. The carbonic anhydrase inhibitor benzolamide (10−4 M, 4 min exposure), which is poorly membrane permeant, had little effect on the rate of cell acidification, indicating that the site of carbonic anhydrase action was intracellular. The activity of Müller cell carbonic anhydrase may help to buffer extracellular CO2 variations in the retina. © 1994 Wiley‐Liss, Inc.
KW - Carbon dioxide regulation
KW - Glial cell
KW - Intracellular pH
KW - Retina
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U2 - 10.1002/glia.440110402
DO - 10.1002/glia.440110402
M3 - Article
C2 - 7960033
AN - SCOPUS:0028485757
SN - 0894-1491
VL - 11
SP - 291
EP - 299
JO - Glia
JF - Glia
IS - 4
ER -