Nicotinic actions on neurones of the central autonomic area in rat spinal cord slices

Abstract

1. Nicotinic responses and actions on excitatory synaptic activity were studied in eighty-four neurones in the region dorsal to the central canal (lamina X) in transverse thoracolumbar spinal cord slices of neonate (P2-P10) rats by using the whole-cell patch-clamp technique. 2. Neurones (n = 15) labelled with Lucifer Yellow, showed the typical morphology of sympathetic preganglionic neurones (SPNs) in the central autonomic area (CA). Unlabelled neurones of comparable morphology were visually identified and recorded. 3. All neurones recorded responded to the nicotinic acetylcholine receptor (nAChR) agonist, DMPP. Under current-clamp conditions, pressure ejections of DMPP depolarized cells and induced the discharge of action potentials. Tetrodotoxin suppressed action potentials but not DMPP-induced depolarization. 4. Under voltage-clamp conditions at a holding potential (Vh) of -50 mV, DMPP induced a transient inward current (which reversed around 0 mV) and an increase in membrane current noise in 50% of the recorded neurones. In the others, DMPP increased membrane current noise without measurable inward current. The current-voltage relationship showed strong inward rectification at holding potentials more positive than 0 mV. 5. In neurones displaying a detectable current response to DMPP, the following agonist rank order potency could be established: DMPP = nicotine > cytisine > ACh. The DMPP response could be blocked by mecamylamine but was insensitive to methyllycaconite. 6. Pressure application of glutamate induced inward currents in all cells tested at a Vh of -50 mV. This response reversed at 10 mV, displayed a region of negative slope conductance at Vh more negative than -30 mV and was partially blocked by CNQX. Pressure application of DMPP transiently increased the amplitude of the glutamate-induced current in six out of nine cells tested. This potentiation persisted in the presence of tetrodotoxin. 7. Forty per cent of the recorded neurones displayed spontaneous excitatory postsynaptic currents (sEPSCs). At a Vh of -50 mV the sEPSCs had a mean amplitude of -19.3 pA and occurred at a frequency below 0.5 Hz. sEPSCs were blocked by CNQX and inverted around 0 mV. Brief application of DMPP increased the discharge frequency of sEPSCs without affecting their kinetics. Additionally, in some cells DMPP increased mean sEPSC amplitude. 8. Focal electrically evoked EPSCs reversed close to 10 mV and were sensitive to CNQX. They occurred with a constant latency, rise time and a mono-exponential decay time. Application of DMPP decreased the percentage of stimulation failures and increased the amplitude of evoked EPSCs, in all cells tested. 9. It is concluded that neurones in the CA, presumed to be SPNs, have functional nAChRs with activation having two distinct effects: firstly, a direct depolarization of the postsynaptic membrane; and secondly, a facilitation of the excitatory transmission onto these cells. This second effect is achieved by an increase of the size of the glutamate-induced current at the postsynaptic level as well as by an enhancement of the presynaptic release of glutamate.