ELECTRORESPONSIVE PROPERTIES AND MEMBRANE-POTENTIAL TRAJECTORIES OF 3TYPES OF INSPIRATORY NEURONS IN THE NEWBORN MOUSE-BRAIN STEM IN-VITRO

1. The electrophysiological properties of inspiratory neurons were stu died in a rhythmically active thick-slice preparation of the newborn m ouse brain stem maintained in vitro. Whole cell patch recordings were performed from 60 inspiratory neurons within the rostral ventrolateral part of the slice with the aim of extending the classification of ins piratory neurons to include analysis of active membrane properties. 2. The slice generated a regular rhythmic motor output recorded as burst of action potentials on a XII nerve root with a peak to peak time of 11.5 +/- 3.4 s and a duration of 483 +/- 54 ms (means +/- SD, n = 50). Based on the electroresponsive properties and membrane potential traj ectories throughout the respiratory cycle, three types of inspiratory neurons could be distinguished. 3. Type-1 neurons were spiking in the interval between the inspiratory potentials (n = 9) or silent with a r esting membrane potential of -48.6 +/- 10.1 mV and an input resistance of 306 +/- 130 M Omega (n = 15). The spike activity between the inspi ratory potentials was burst-like with spikes riding on top of an under lying depolarization (n = 11) or regular with no evidence of bursting (n = 12). Hyperpolarization of the neurons below threshold for spike i nitiation did not reveal any underlying phasic synaptic activity, that could explain the bursting behavior. 4. Type-1 neurons showed delayed excitation after hyperpolarizing square current pulses or when the ne urons were depolarized from a hyperpolarized level. This membrane beha vior resembles the response seen in other CNS neurons expressing an I- A. The response to 1-s long depolarizing pulses with a large current s trength showed signs of activation of an active depolarizing membrane response leading to a transient reduction in the spike amplitude. The relationship between the membrane potential and the amplitude of squar e current pulses (V-m-I) showed a small upward rectification below -70 mV, and spike adaptation throughout a 1-s pulse had a largely linear time course. 5. Type-1 neurons depolarized and started to fire spikes 398 +/- 102 ms (n = 20) before the upstroke of the integrated XII nerv e discharge. The inspiratory potential was followed by fast hyperpolar ization, a short fast-repolarizing phase (1,040 +/- 102 ms, n = 5) and a longer slow-repolarizing phase (lasting until the next inspiratory discharge). 6. Type-2 neurons were spiking in the interval between the inspiratory potentials with no evidence of bursting behavior and had an input resistance of 296 +/- 212 M Omega (n = 26), The response to h yperpolarizing pulses revealed an initial sag and postinhibitory rebou nd depolarization. This membrane behavior resembles the response seen in other CNS neurons expressing an I-h. The V-m-I relationship was lin ear at depolarized potentials and showed a marked upward rectification below -69 mV. Spike trains elicited by 1-s long pulses showed a prono unced early and late adaptation. 7. Type-2 neurons depolarized and sta rted to fire spikes 171 +/- 87 ms (n = 23) before the upstroke of the integrated XII nerve discharge. The inspiratory potential had a variab le amplitude from cell to cell and was followed by a short hyperpolari zation in the cells displaying a large amplitude inspiratory potential . 8. Type-3 neurons were spiking in the interval between the inspirato ry potentials with no evidence of bursting behavior (n = 5) or silent with a resting membrane potential of -50.4 +/- 2.7 mV and an input res istance of 126 +/- 34 M Omega (n = 10). The response to current pulses revealed a low-threshold depolarization elicited by depolarizing puls es from a hyperpolarized level or after hyperpolarizing pulses from re st. This membrane behavior resembles the response seen in other CNS ne urons expressing a low-voltage-activated Ca2+ current. The V-m-I relat ionship showed a small upward rectification below -70 mV and the spike train throughout a 1-s pulse showed a pronounced early and late adapt ation. 9. Type-3 neurons depolarized 100 +/- 40 ms (n = 9) before the upstroke of the integrated XII nerve discharge. The inspiratory potent ial was followed by a small amplitude afterdepolarization at resting m embrane potential. 10. In conclusion, three types of inspiratory neuro ns in the newborn mouse ventrolateral brain stem display a number of a ctive membrane responses, which seem to shape the membrane potential t rajectory before, during, and after the inspiratory potentials. The di fferential timing of synaptic activity preceding the XII nerve dischar ge demonstrates a sequence of activation in relation to the inspirator y burst, where type-1 neurons are activated well before type-2 neurons , which are activated before type-3 neurons suggesting that type-1 neu rons may be more closely involved in the generation of the respiratory rhythm.