Functional characteristics of H+-dependent nicotinate transport in primary cultures of astrocytes from rat cerebral cortex

In the present study, we report the characteristics of H+-coupled nicotinate transport in primary cultures of astrocytes from rat cerebral cortex. The [3H]nicotinate transport in rat astrocytes increased up to a pH 5.5. The nicotinic acid uptake at pH 6.0 was both energy-dependent and saturable with a Michaelis constant (Kt) of 2.8 ± 0.4 mM and the maximal uptake rate (Vmax) of 31 ± 3.2 nmol/mg protein/10 min. This process was reduced by a protonophore, carbonylcyanide p-trifluoromethoxyphenylhydrazone, and a typical monocarboxylate transporter (MCT) inhibitor, α-cyano-4-hydroxycinnamic acid, suggesting that nicotinate uptake by rat astrocytes is mediated by H+-coupled monocarboxylate transport system. [3H]Nicotinate transport in rat astrocytes was significantly inhibited by various monocarboxylic acids such as l-lactic acid and pyruvic acid with a relatively low affinity (Ki > 10 mM). On the other hand, the uptake process of l-lactic acid was also saturable with a high-affinity component (Kt = 0.27 mM) and a low-affinity component (Kt = 35.9 mM). Reverse transcription-PCR and Western blot analyses revealed that three MCT subtypes, MCT1/Slc16a1, MCT2/Slc16a7, and MCT4/Slc16a3, were expressed in these cells. Because l-lactate reduced to 67% of the nicotinate uptake even at 10 mM, it is unlikely that nicotinate uptake in rat astrocytes is mediated by MCT1 and/or MCT2. These results provide biochemical evidence of a H+-coupled and saturable transport system, presumed to be a low-affinity monocarboxylate transporter MCT4 or other unknown H+-coupled monocarboxylate transport system, for nicotinate in rat cerebrocortical astrocytes.