Electrophysiological Characterization of Non-NMDA Glutamate Receptors on Cultured Intermediate Lobe Cells of the Rat Pituitary

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Abstract
Glutamate is the major excitatory neurotransmitter in the central nervous system, yet little is known about its actions on endocrine cells. We have investigated the membrane effects of glutamate in cultured neonatal rat pituitary intermediate lobe (IL) cells using the whole-cell configuration of the patch-clamp technique. In a standard Na+-based extracellular solution, glutamate failed to induce a detectable membrane current at a holding potential (HP) of –60 mV (n = 40). However, when cyclothiazide (50 µM), a benzothiazide that blocks desensitization of α-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazole-propanoic acid (AMPA)-type receptors, was added to the extracellular solution, glutamate (0.5–1 mM) induced an inward current at a HP of –60 mV in 65% of the cells tested (n = 72). This response was usually small in amplitude (mean amplitude: 28.6 ± 37.5 pA, n = 47). The glutamate-induced current reversed polarity close to 0 mV and was reversibly blocked when extracellular Na+ was replaced by the impermeant cation N-methyl-D-glucamine, suggesting that this current was a nonselective cation current. The response to glutamate (1 mM) was reproduced by AMPA (50 µM), kainate (200 µM), and quisqualate (200 µM). N-Methyl-D-aspartate (NMDA, 100 µM) in the presence of 10 µM glycine did not induce any membrane current in cells responding to glutamate (n = 8). The non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (10 µM) reversibly inhibited the response to glutamate (0.5 mM) by 85 ± 14% (n = 7), whereas D(–)-2-amino-5-phosphono-pentanoic acid (20 µM), an antagonist of NMDA receptors, had no effect on the glutamate-induced current (n = 3). Moreover, we show that although the amplitude of the glutamate currents was small, the latter induced large (30-mV) membrane depolarizations and triggered the firing of action potentials. Taken together, our results indicate that neonatal rat IL cells possess AMPA-type glutamate receptors that could possibly underlie a fast excitatory glutamatergic synaptic input to these cells.

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