Regulation of pH in rat brain synaptosomes. II. Role of Cl-.
1. We have previously shown that rat brain synaptosomes exhibit a very dynamic Na+/H+ exchanger. We have also observed that although synaptosomes lack HCO3(-)-based transport mechanisms, they do respond with changes in pHi upon Clo- removal. 2. Here we show that when synaptosomes are transferred from Ringer solution (RS) to Cl(-)-free RS, there is a cytosolic alkalinization of approximately 0.22 pH units. This phenomenon is DIDS (4,4'-diisothiocyanostilbene-3,3'-disulfonic acid) inhibitable. The alkalinization is completely reversed when Cl- ions are reintroduced. The presence of HCO3- or Ca2+ does not modify the response to Cl(-)-removal or replenishment. 3. In acid-loading experiments, the initial rate of pHi recovery is higher in Cl(-)-free RS than in RS. The final resting pHi after the recovery in Cl(-)-free RS is approximately 0.22 pH units higher than that obtained in media containing Cl-. The magnitude of the NaOAc-induced acidification is 2.5-fold larger in the presence than in the absence of Cl-. Similar results are obtained in the presence of HCO3-. 4. These data suggest that H+ movements may be coupled to Cl- movements. To study this possibility further, we developed a technique to simultaneously measure H+ and Cl- by using the fluorescence of 5' (and 6')-carboxy-10-dimethylamino-3-hydroxy-spyro-[7H benzo[c]xanthene- 7,1'(3'H)-isobenzofuran]3'-one (SNARF-1) and MQAE [N-(6-methoxyquinolyl)acetoxy ester], respectively. 5. Our results indicated that the steady-state [Cl-]i in synaptosomes is approximately 56 mM, thus indicating that Cli- is not passively distributed.(ABSTRACT TRUNCATED AT 250 WORDS)
Martínez-Zaguilán
R
R
Department of Biochemistry, University of Arizona, Tucson 85724.
Gillies
R J
RJ
Sánchez-Armass
S
S
D000136:Acid-Base Equilibrium / Q000502:physiology*; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism; D001921:Brain / Q000502:physiology*; D002352:Carrier Proteins / Q000502:physiology; D002712:Chlorides / Q000502:physiology*; D046508:Culture Techniques; D006863:Hydrogen-Ion Concentration; D051381:Rats; D013574:Synaptosomes / Q000502:physiology*
D001639:Bicarbonates; D002352:Carrier Proteins; D002712:Chlorides