P19 cells differentiate into glutamatergic and glutamate-responsive neurons in vitro

• Published in: Neuroscience Vol. 80; no. 2; pp. 487 - 499
• Main Authors: MacPherson, P.A, Jones, S, Pawson, P.A, Marshall, K.C, McBurney, M.W
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 01.09.1997; Elsevier
• Subjects: Animals; Biological and medical sciences; Blotting, Northern; Cell Differentiation; Cell Line; Electrophysiology; embryonal carcinoma; Fluorescent Antibody Technique, Direct; Fundamental and applied biological sciences. Psychology; gamma-Aminobutyric Acid - physiology; Glutamic Acid - physiology; Immunohistochemistry; Isolated neuron and nerve. Neuroglia; Neurons - physiology; Neurons - ultrastructure; neurotransmitter; Patch-Clamp Techniques; Rats; receptor; Receptors, Glutamate - metabolism; Receptors, Glutamate - physiology; retinoic acid; Synapses - physiology; Synapses - ultrastructure; Vertebrates: nervous system and sense organs; NeuN, neuron-specific nucleoprotein; NMDAR, NMDA receptor; HEPES, N-2-hydroxyethylpiperazine- N′-2-ethanesulphonic acid; MK-801, dizocilpine maleate; FITC, fluorescein isothiocyanate; embryonal carcinoma; EGTA, ethyleneglycolbis(aminoethylether)tetra-acetate; GluR, glutamate receptor; TTX, tetrodotoxin; retinoic acid; receptor; PBS, phosphate-buffered saline; AMPA, amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; PCR, polymerase chain reaction; neurotransmitter; NMDA, N-methyl- d-aspartate; Cell line; Neuron; Excitatory aminoacid; Neurotransmitter; Glutamate receptor; Retinoids; Glutamate; In vitro
• ISSN: 0306-4522; 1873-7544
• DOI: 10.1016/S0306-4522(97)00102-4

The neurotransmitter l-glutamate has been associated with a number of developmental events within the central nervous system including synaptogenesis and the refinement of topographically ordered neural maps. As a model for studying such events at the molecular level, we have examined the expression of glutamate and glutamate receptors in neurons that develop from P19 cells in response to retinoids. We report here that many P19-derived neurons do contain glutamate in secretory vesicles and that this glutamate appears to function as a neurotransmitter. The neurotransmitter GABA is also present in these cultures and both glutamate and GABA appeared to co-localize in some neuronal processes. Both neurotransmitters were released from the neurons in response to membrane depolarization. These neurons also express various glutamate receptor subunits including GluR1, GluR4 and NMDAR1 as detected by immunological methods. Using whole-cell patch-clamping, we have recorded spontaneous postsynaptic potentials which increase in both amplitude and frequency with time in culture and which are sensitive to the glutamate antagonist kynurenic acid. Thus, P19-derived neurons mature in culture and form electrically active neural networks involving glutamate and glutamate receptors.