EXCITATION OF SYMPATHETIC PREGANGLIONIC NEURONS VIA METABOTROPIC EXCITATORY AMINO-ACID RECEPTORS

The role of excitatory amino acid metabotropic receptors in the regulation of excitability of sympathetic preganglionic neurons was investigated. This study used both conventional intracellular and whole-cell patch clamp techniques to record from sympathetic preganglionic neurons in transverse spinal cord slices of the rat (9-21 days old) The metabotropic receptor agonists (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) (10-200 mu M, superfused for 2-60 s) and quisqualate (1-50 mu M, superfused for 2-60 s) induced concentration-dependent depolarizing responses which did not desensitize. These responses were unaffected by the glutamate ionotropic receptor antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10-50 mu M), 6,7-dinitroquinoxaline-2,3-dione (DNQX, 10 mu M), dizocilpine (MK-801, 10-40 mu M), 3-[(R)-2-carboxy-piperazin-4-yl]-propyl-1-phosphonic acid (D-CPP, 10-50 mu M) and DL-2-amino-5-phosphonovaleric acid (DL-AP5, 20-100 mu M). Depolarizing responses to 1S,3R-ACPD and quisqualate were unaffected by L-2-amino-3-phosphonopropionic acid (L-AP3, 30 mu M-1 mM) and L-2-amino-4-phosphonobutanoic acid (L-AP4, 100 mu M-1 mM). The responses to 1S,3R-ACPD and quisqualate were reduced by including the G-protein blocker GDP-beta-S (400 mu M) in the patch pipette solution by 77 +/- 2% (mean +/- S.E) of control (n = 3), suggesting that these agonists activate a G-protein-coupled receptor. Metabotropic receptor-mediated responses were maintained in the presence of tetrodotoxin (500 nM), progressively reduced with increased membrane hyperpolarization to around -95 mV and associated with either an increase of 16.5 +/- 2.8% (data from four neurons) in the majority of neurons (n = 22 of 34) or no measurable change (n = 12) in neuronal input resistance. These data suggest that the agonists exert a direct action on sympathetic preganglionic neurons involving a reduction in one or more membrane conductances. 1S,3R-ACPD and quisqualate had several effects on sympathetic preganglionic neuron membrane properties including: inhibition of a slow apamin-insensitive component of the afterhyperpolarization; a reduction in spike frequency adaptation leading to increases in firing frequency from 6.4 +/- 2.8 Hz in control experiments up to 14.7 +/- 3.0 Hz (n = 6 neurons) in the presence of a metabotropic receptor agonist; a broadening of the action potential by 37.5 +/- 6.4% (n = 6 neurons) of control. These observations suggest that the metabotropic receptor-mediated depolarization is due, at least in part, to the reduction of potassium conductances involved in the spike afterhyperpolarisation potential. In quiescent sympathetic preganglionic neurons, 1S,3R-ACPD and quisqualate induced rhythmic oscillations in membrane potential. This effect could be irreversible with oscillations persisting for several hours post-induction. These oscillations could give rise to sustained, rhythmic activity characterized by burst firing or regular single spike discharge. The oscillations were similar to those observed by us to occur spontaneously in some sympathetic preganglionic neurons. The amplitude and frequency of oscillations varied considerably in the same neuron, due to different rhythms being superimposed on top of one another, and between different sympathetic preganglionic neurons. The mean peak amplitude and frequency of oscillations observed with patch electrodes at room temperature was 9.6 +/- 2.3 mV and 0.31 +/- 0.05 Hz (data pooled from six sympathetic preganglionic neurons) respectively. These effects were not sensitive to ionotropic receptor antagonists and were not mimicked by the ionotropic receptor agonists AMPA or kainate, suggesting selective activation of metabotropic receptors is required to induce oscillatory activity in some previously silent sympathetic preganglionic neurons. As oscillations are thought to reflect electrotonic coupling between sympathetic preganglionic neurons these observations suggest metabotropic receptors may have an important role to play in synchronizing the electrical activity of groups of sympathetic preganglionic neurons. The results suggest that sympathetic preganglionic neurons possess functional metabotropic excitatory amino acids receptors whose activation increases the excitability of these neurons by reducing potassium conductances. In addition, activation of metabotropic receptors can lead to the induction of rhythmic oscillations in membrane potential and long-term changes in the excitability of these neurons.