L-proline activates glutamate and glycine receptors in cultured rat dorsal horn neurons.
The pharmacological actions of L-proline on excitatory and inhibitory amino acid receptors have been characterized under voltage-clamp conditions, using cultured dissociated neurons from the dorsal horn of the rat spinal cord. At a holding potential of -62 mV, millimolar concentrations of L-proline elicited an inward current that was partially antagonized by D-(-)-2-amino-5-phosphonopentanoic acid (APV), 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), and strychnine and was virtually abolished (97% block) by a combination of all three antagonists. Currents evoked by D-proline were abolished by strychnine alone. APV-, CNQX-, and strychnine-sensitive components of L-proline-evoked currents were isolated using various combinations of the three antagonists. These currents were identical to currents elicited by N-methyl-D-aspartate (NMDA), kainate, and glycine, respectively, with respect to antagonist specificity, reversal potential, and ionic permeability. The APV- and strychnine-sensitive currents also showed a time dependence similar to that of the currents elicited by NMDA and glycine. EC50 values could not be calculated, because the response did not saturate within the tested range of L-proline concentrations (0.3-50 mM). Estimates of relative potency were obtained, however, by comparison with responses elicited by selective agonists. The APV-sensitive, CNQX-sensitive, and strychnine-sensitive currents evoked by 10 mM L-proline were comparable in size to currents elicited by 15 microM NMDA, 5 microM kainate, and 30 microM glycine, respectively. L-Proline was found to elicit an increase in intracellular [Ca2+] that was dependent upon Ca2+ entry into the cell. These Ca2+ responses were enhanced by strychnine and partially antagonized by APV, CNQX, or Mg2+. Our results using dorsal horn neurons grown in culture indicate that L-proline is a weak agonist at strychnine-sensitive glycine receptors and at both NMDA and non-NMDA glutamate receptors. These observations should help in interpreting the confusing array of L-proline actions that have been described using more intact nervous system preparations. Furthermore, the ability of L-proline to stimulate Ca2+ entry after activation of excitatory amino acid receptors implicates L-proline as a potential endogenous excitotoxin.
Department of Physiology and Cellular Biophysics, Columbia University, New York, New York 10032.
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Henzi
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Reichling
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Helm
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MacDermott
D015763:2-Amino-5-phosphonovalerate / Q000494:pharmacology; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D005260:Female; D005971:Glutamates / Q000378:metabolism*; D005998:Glycine / Q000378:metabolism*; D007608:Kainic Acid / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D016202:N-Methylaspartate / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D011247:Pregnancy; D011392:Proline / Q000494:pharmacology*; D011810:Quinoxalines / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D017470:Receptors, Glutamate; D018009:Receptors, Glycine; D017981:Receptors, Neurotransmitter / Q000187:drug effects*; D013116:Spinal Cord / Q000166:cytology; D013331:Strychnine / Q000494:pharmacology
D005971:Glutamates; D011810:Quinoxalines; D017470:Receptors, Glutamate; D018009:Receptors, Glycine; D017981:Receptors, Neurotransmitter; D016202:N-Methylaspartate; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D015763:2-Amino-5-phosphonovalerate; D011392:Proline; D013331:Strychnine; D007608:Kainic Acid; D002118:Calcium; D005998:Glycine