The regional distribution of spinal cord networks producing locomotor-like, as well as non-locomotor-like, activity was studied with the use of an in vitro neonatal rat preparation. Rhythmic activity was induced by bath application of either serotonin (5-HT), acetylcholine (ACh), N-methyl-D,L-aspartate (NMA), or combined 5-HT/NMA, and was monitored via hindlimb flexor (peroneal) and extensor (tibial) electroneurograms (ENGs) or ventral root recordings. In some experiments, synchronous patterns were produced by the addition of inhibitory amino acid (IAA) receptor antagonists. Selective application of 5-HT to cervical and thoracic cord regions induced rhythmic activity in these segments but failed to evoke hindlimb ENG discharge. Exposure of the isolated lumbar region to 5-HT produced tonic activity only. Application of 5-HT to the whole cord produced locomotor-like activity in hindlimb ENGs that persisted after midsagittal section of the spinal cord from the conus to the thoracolumbar junction. In other experiments, transverse hemisection of the rostral lumbar cord during whole cord exposure to 5-HT abolished rhythmic activity in ipsilateral hindlimb ENGs, suggesting that under these conditions rhythmic activity on one side of the lumbar cord was insufficient to maintain rhythmic activity on the contralateral side. Selective application of NMA or ACh to cervical and/or thoracic cord regions evoked rhythmic activity in these supralumbar segments, as well as rhythmic, but non-locomotor-like, activity in the lumbar region. In contrast to the effect of 5-HT, both NMA and ACh evoked rhythmic activity when applied solely to the lumbar region, and the side-to-side alternation produced by whole cord ACh application was uncoupled by midsagittal lesions of the lumbar region. In the presence of IAA antagonists, the side-to-side coupling of bilaterally synchronous rhythms was maintained despite extensive midsagittal lesions leaving all but one or two segments of either cervical, thoracic, or lumbar cord bilaterally intact, and rhythmic activity could be maintained even in single isolated hemisegments. The effects of 5-HT/NMA were similar to those observed with the use of 5-HT alone, although 5-HT/NMA induced rhythmic activity in hindlimb ENGs when applied selectively to supralumbar regions. The results suggest that 1) a 5-HT-sensitive oscillatory network, capable of producing a locomotor-like pattern of activity, is distributed throughout the supralumbar region of the spinal cord and mediates descending rhythmic drive to lumbar motor centers; 2) NMA- and ACh-sensitive rhythmogenic elements are distributed throughout the spinal cord, including the lumbar region; and 3) the spinal cord contains an extensive propriospinal network of reciprocal inhibitory and excitatory connections characterized by redundantly organized side-to-side projections.

1. The role of inhibitory amino acid transmission in the coordination and generation of rhythmic motor activity was examined with the use of an in vitro neonatal rat spinal cord preparation. Before adding gamma-aminobutyric acid (GABA) or glycine receptor agonists and antagonists, rhythmic motor activity was induced by bath application of acetylcholine (ACh), N-methyl-D,L-aspartate (NMA), or serotonin (5-HT) while monitoring bilateral ankle flexor and extensor electroneurograms (ENGs). The timing of rhythmic flexor and extensor discharge was consistent with that seen during overground locomotion in 27% of 84 bath applications of these substances (n = 65 preparations). 2. Subsequent addition of the GABAA receptor agonist muscimol, the GABAB receptor agonist baclofen, or glycine, abolished rhythmic activity in 95% of the tested applications. 3. GABAB receptor blockade did not disrupt alternating patterns of ENG discharge. However, addition of the GABAA receptor antagonist bicuculline, or the glycine receptor antagonist strychnine, transformed alternating flexor-extensor and left-right activity into patterns characterized by bilaterally synchronous rhythmic activation of all hindlimb ENGs. The onset of individual ENG bursts was more abrupt following bicuculline or strychnine. Strychnine also synchronized high-frequency (4-8 Hz) packets of rhythmic discharge within ENG bursts. 4. Some preparations developed synchronous, but unstable, rhythmic activity in the presence of bicuculline or strychnine alone. However, NMA, 5-HT, or ACh was usually required in addition to these antagonists to promote sustained rhythmic activity.(ABSTRACT TRUNCATED AT 250 WORDS)

No abstract

We recently showed that propriospinal neurons contribute to bulbospinal activation of locomotor networks in the in vitro neonatal rat brainstem-spinal cord preparation. In the present study, we examined whether propriospinal neurons alone, in the absence of long direct bulbospinal transmission to the lumbar cord, can successfully mediate brainstem activation of the locomotor network. In the presence of staggered bilateral spinal cord hemisections, the brainstem was stimulated electrically while recording from lumbar ventral roots. The rostral hemisection was located between C1 and T3 and the contralateral caudal hemisection was located between T5 and mid-L1. Locomotor-like activity was evoked in 27% of the preparations, which included experiments with staggered hemisections placed only two segments apart. There was no relation between the likelihood of developing locomotor-like activity and the distance separating the two hemisections or specific level of the hemisections. In some experiments, where brainstem stimulation alone was ineffective, neurochemical excitation of propriospinal neurons (using 5-HT and NMDA) at concentrations subthreshold for producing locomotor-like activity, promoted locomotor-like activity in conjunction with brainstem stimulation. In other experiments, involving neither brainstem stimulation nor cord hemisections, the excitability of propriospinal neurons in the cervical and/or thoracic region was selectively enhanced by bath application of 5-HT and NMDA or elevation of bath K + concentration. These manipulations produced locomotor-like activity in the lumbar region. In total, the results suggest that propriospinal neurons are sufficient for transmission of descending locomotor command signals. This observation has implications for regeneration strategies aimed at restoration of locomotor function after spinal cord injury.

The effect of serotonin (5-HT) receptor blockade on rhythmic network activity and on N-methyl--aspartate (NMDA) receptor-induced membrane voltage oscillations was examined using an in vitro neonatal rat spinal cord preparation. Pharmacologically induced rhythmic hindlimb activity, monitored via flexor and extensor electroneurograms or ventral root recordings, was abolished by 5-HT receptor antagonists. Intrinsic motoneuronal voltage oscillations, induced by NMDA in the presence of tetrodotoxin (TTX), either were abolished completely or transformed to long-lasting voltage shifts by 5-HT receptor antagonists. Conversely, 5-HT application facilitated the expression of NMDA-receptor-mediated rhythmic voltage oscillations. The results suggest that an interplay between 5-HT and NMDA receptor actions may be critical for the production of rhythmic motor behavior in the mammalian spinal cord, both at the network and single cell level.