Neurotransmitter release from growth cones of rat dorsal root ganglion neurons in culture

Abstract

Growing neurites of rat dorsal root ganglion neurons in culture formed growth cones at the tips. Possible release of glutamate from these growth cones was investigated by using a whole-cell patch-clamp recording from an acutely dissociated hippocampal neuron containing glutamate receptors. The hippocampal neuron was placed in contact to various regions of the dorsal root ganglion neurons. Inward currents were recorded from the hippocampal neuron positioned on the growth cones of the dorsal root ganglion neurons (diameter, 12–16 μm) in response to the dorsal root ganglion cell body stimulation. The inward currents were associated with an increase in membrane conductance, and the reversal potential was estimated at −6.5 mV ( n=8). The inward currents were blocked by 6-cyano-7-nitroquinoxaline (10 μM), but not blocked by 2-amino-5-phosphonovaleric acid (50 μM) and bicuculline (10 μM). The inward currents were abolished by tetrodotoxin (1 μM), EGTA-buffered Ca 2+-free external solution or ω-agatoxin IVA (300 nM), and were inhibited by ω-conotoxin GVIA (3 μM), but were not affected by nicardipine (10 μM). Intracellular calcium ion concentration ([Ca 2+] i) in growth cones of the dorsal root ganglion neurons increased in response to dorsal root ganglion cell body stimulation, whereas the elevation of [Ca 2+] i was not observed either in the presence of tetrodotoxin (1 μM) or in a Ca 2+-free external solution. These results indicate that the inward currents were evoked by glutamate released from the growth cones via a Ca 2+-dependent process, and suggest that the growth cones are already endowed with much of the machinery for neurotransmitter release, even before making a structure for synaptic transmission.