L-TRANs-Pyrrolidine-2,4-dicarboxylate and cis-1-aminocyclobutane-1, 3-dicarboxylate behave as transportable, competitive inhibitors of the high-affinity glutamate transporters

The ability of two conformationally restricted analogues of L-glutamate to function as nontransportable inhibitors of plasma membrane L-glutamate transport was investigated in primary cultures of cerebellar granule cells and cortical astrocytes. L-trans-Pyrrolidine-2,4-dicarboxylic acid (L-transPDC) and cis-1-aminocyclobutane-1,3-dicarboxylic acid (cis-ACBD) behaved as linear competitive inhibitors of the uptake of d-[³H]aspartate (used as a non-metabolizable analogue of L-glutamate) exhibiting K_i values between 40 and 145 μm; L-trans-PDC being the more potent inhibitor in each preparation. However, both L-trans-PDC and cis-ACBD, over a concentration range of 1 μM-5 mM, dose-dependently stimulated the release of exogenously supplied d-[³H]aspartate from granule cells maintained in a continuous superfusion system. The stimulated release was independent of extracellular calcium ions; essentially superimposable dose-response profiles being obtained in the absence and presence of 1.3 mM CaCl₂ and yielding ec₅₀ values of 16-25 μM and 180-220 μM for L-trans-PDC and cis-ACBD, respectively. Stimulated release of d-[³H]aspartate was unaffected by either 300 μm d-(-)-2-amino-5-phosphonopentanoi acid [d-APV; a selective antagonist of the N-methyl-d-aspartate (NMDA) receptor] or by 25 μM 6-cyano-7-nitroquinoxaline-2,3-dione [CNQX; a selective antagonist of the α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor]. The release of d-[³H] aspartate following stimulation by either L-trans-PDC or cis-ACBD was however markedly attenuated following substitution in the superfusion medium of sodium ions by choline ions. Taken together, these results support an action of L-trans-PDC and cis-ACBD consistent with that of being competitive substrates rather than non-transportable blockers of the plasma membrane L-glutamate uptake system.