Ventral Root Discharges Research Articles

Growth restriction induced by chronic prenatal hypoxia affects breathing rhythm and its pontine catecholaminergic modulation

Impaired transplacental supply of oxygen leads to intrauterine growth restriction, one of the most important causes of perinatal mortality and respiratory morbidity. Breathing rhythm depends on the central respiratory network modulated by catecholamines. We investigated the impact of growth restriction, using prenatal hypoxia, on respiratory frequency, on central respiratory-like rhythm, and on its catecholaminergic modulation after birth. At birth, respiratory frequency was increased and confirmed in en bloc medullary preparations, where the frequency of the fourth cervical (C4) ventral root discharge was increased, and in slice preparations containing the pre-Bötzinger complex with an increased inspiratory rhythm. The inhibition of C4 burst discharge observed in pontomedullary preparations was stronger in the growth-restricted group. These results cannot be directly linked by the tyrosine hydroxylase activity increase of A1/C1 and A2/C2 cell groups in the medulla since blockade of α1- and α2-adrenergic receptors did not abolish the difference between both groups. However, in pontomedullary preparations, the stronger inhibition of C4 burst discharge is probably supported by an increased inhibition of A5, a respiratory rhythm inhibitor pontine group of neurons displaying increased tyrosine hydroxylase activity, because blockade of α2-adrenergic receptors abolished the difference between the two groups. Taken together, these results indicate that growth restriction leads to a perturbation of the breathing frequency, which finds, at least in part, its origin in the modification of catecholaminergic modulation of the central breathing network.

Facilitation of ipsilateral actions of corticospinal tract neurons on feline motoneurons by transcranial direct current stimulation.

Ipsilateral actions of pyramidal tract (PT) neurons are weak but may, if strengthened, compensate for deficient crossed PT actions following brain damage. The purpose of the present study was to examine whether transcranial direct current stimulation (tDCS) can strengthen ipsilateral PT (iPT) actions; in particular, those relayed by reticulospinal neurons co-excited by axon collaterals of fibres descending in the iPT and contralateral PT (coPT) and of reticulospinal neurons descending in the medial longitudinal fascicle (MLF). The effects of tDCS were assessed in acute experiments on deeply anaesthetized cats by comparing postsynaptic potentials evoked in hindlimb motoneurons and discharges recorded from their axons in a ventral root, before, during and after tDCS. tDCS was consistently found to facilitate joint actions of the iPT and coPT, especially when they were stimulated together with the MLF. Both excitatory postsynaptic potentials and inhibitory postsynaptic potentials evoked in motoneurons and the ensuing ventral root discharges were facilitated, even though the facilitatory effects of tDCS were not sufficient for activation of motoneurons by iPT neurons alone. Facilitation outlasted single tDCS periods by at least a few minutes, and the effects evoked by repeated tDCS by up to 2h. The results of this study thus indicate that tDCS may increase the contribution of iPT actions to the recovery of motor functions after injuries to coPT neurons, and thereby assist rehabilitation, provided that corticoreticular and reticulospinal connections are preserved.

Contribution of TRPV1 Receptor–Expressing Fibers to Spinal Ventral Root After-Discharges and Mechanical Hyperalgesia in a Spared Nerve Injury (SNI) Rat Model

Neuropathic pain induces allodynia and hyperalgesia. In the spared nerve injury (SNI) model, marked mechanical hyperalgesia is manifested as prolongation of the duration of paw withdrawal after pin stimulation. We have previously reported that spinal ventral root discharges (after-discharges) after cessation of noxious mechanical stimulation applied to the corresponding hindpaw were prolonged in anesthetized spinalized rats. Since these after-discharges occurred through transient receptor potential (TRP) V1–positive fibers, these fibers could contribute to mechanical hyperalgesia. Therefore, we examined whether selective deletion of TRPV1-positive fibers by resiniferatoxin, an ultrapotent TRPV1 agonist, would affect the behavioral changes and ventral root discharges in SNI rats. Mechanical allodynia in the von Frey test, mechanical hyperalgesia after pin stimulation, and enhancement of ventral root discharges, but not thermal hyperalgesia in the plantar test, appeared in Wistar rats with SNI. Mechanical hyperalgesia was abolished by treatment with resiniferatoxin, whereas mechanical allodynia was not affected. Moreover, resiniferatoxin eliminated after-discharges completely. These results show that TRPV1-positive fibers do not participate in the mechanical allodynia caused by sensitization of Aβ-fibers, but contribute to the enhancement of after-discharges and mechanical hyperalgesia following SNI. It is suggested that the mechanisms responsible for generating mechanical allodynia differ from those for prolongation of mechanical hyperalgesia.

N- and L-Type Voltage-Dependent Ca2+ Channels Contribute to the Generation of After-Discharges in the Spinal Ventral Root After Cessation of Noxious Mechanical Stimulation

Voltage-dependent Ca2+ channels (VDCCs) play a crucial role in the spinal pain transduction. We previously reported that nociceptive mechanical stimuli to the rat hindpaw evoked two types of ventral root discharges that increased during stimulation (during-discharges) and after cessation of stimulation (after-discharges). To explore the involvement of VDCCs in these ventral root discharges, several VDCC blockers were applied directly to the surface of the spinal cord. Spinalized rats were laminectomized. The fifth lumbar ventral root was sectioned and used for multi-unit efferent discharges recording. An agar pool was constructed on the first lumbar vertebra for drug application. Ethosuximide (a T-type VDCC blocker) had no effect on ventral root discharges. ω-Conotoxin GVIA (an N-type VDCC blocker) preferentially suppressed after-discharges. ω-Agatoxin IVA (a P/Q-type VDCC blocker), diltiazem, and verapamil (L-type VDCC blockers) nonselectively depressed both during- and after-discharges. The more selective L-type VDCC blocker nicardipine depressed only after-discharges and the depression was exhibited when nicardipine was microinjected into the dorsal horn, but not into the ventral horn. These findings suggested that N- and L-type VDCCs in the dorsal horn were involved in the generation of afterdischarges and these blockers might be useful for treatment of persistent pain that involves the spinal pathway.

Primary Afferent Terminals Acting as Excitatory Interneurons Contribute to Spontaneous Motor Activities in the Immature Spinal Cord

Patterned, spontaneous activity plays a critical role in the development of neuronal networks. A robust spontaneous activity is observed in vitro in spinal cord preparations isolated from immature rats. The rhythmic ventral root discharges rely mainly on the depolarizing/excitatory action of GABA and glycine early during development, whereas at later stages glutamate drive is primarily responsible for the rhythmic activity and GABA/glycine are thought to play an inhibitory role. However, rhythmic discharges mediated by the activation of GABA(A) receptors are recorded from dorsal roots (DRs). In the present study, we used the in vitro spinal cord preparation of neonatal rats to identify the relationship between discharges that are conducted antidromically along DRs and the spontaneous activity recorded from lumbar motoneurons. We show that discharges in DRs precede those in ventral roots and that primary afferent depolarizations (PADs) start earlier than EPSPs in motoneurons. EPSP-triggered averaging revealed that the action potentials propagate not only antidromically in the DR but also centrally and trigger EPSPs in motoneurons. Potentiating GABAergic antidromic discharges by diazepam increased the EPSPs recorded from motoneurons; conversely, blocking DR bursts markedly reduced these EPSPs. High intracellular concentrations of chloride are maintained in primary afferent terminals by the sodium-potassium-chloride cotransporter NKCC1. Blocking these cotransporters by bumetanide decreased both dorsal and ventral root discharges. We conclude that primary afferent fibers act as excitatory interneurons and that GABA, through PADs reaching firing threshold, is still playing a key role in promoting spontaneous activity in neonates.

A novel neural substrate for the transformation of olfactory inputs into motor output.

Author SummaryAnimal behaviors, including locomotion, can be driven by olfactory cues, such as pheromones or food sources. The neural substrate (neuroanatomical connections and physiological signals) that permits the transformation of olfactory inputs into locomotor responses is still unknown in vertebrates. In the present study, we identify such a neural substrate in the lamprey. Here, olfactory signals from the outside world are transmitted to the reticulospinal neurons in the lower brainstem, which provide the descending locomotor command to the spinal cord. We found that this circuit originates in the medial portion of the olfactory bulb and that connections are made in the posterior tuberculum, a ventral diencephalic structure. These inputs are then conveyed to the mesencephalic locomotor region, known to project extensively to brainstem reticulospinal neurons and thereby activate locomotion. Our results illuminate a specific dedicated neural substrate in the brain of lampreys that underlies olfactory-motor responses, which is activated by both food-related or pheromonal olfactory cues. It will be of interest to determine whether such a pathway is preserved in all vertebrates.

Development of respiratory function in perinatal ontogenesis

In development of respiratory function in rats, mice, and other representatives of placental animals there exists the general plan of formation of rhythm: from single contraction of respiratory musculature to formation of bursts and complexes alternating periodically with pauses and apnea intervals and subsequent rhythm stabilization. These peculiarities are closely connected with the states of sleep and consciousness. A concept is put forward about a certain sequence of functional maturation and ways of regulation of activity of the respiratory rhythm central pacemaker. At the first stage the autogenic rhythmical activity is determined by pacemaker properties of a part of neurons of the medulla rostral ventrolateral part. It is not ruled out that the first respiratory discharges in spinal cord ventral roots might have been a manifestation of the nervous network rhythmogenic properties. The direct sensitivity of central neurons to chemical composition if the medium and to some neutomodulators serves as the first regulatory mechanism. Somewhat later, inhibitory control is established from supramedullary structures, with an increase of role of peripheral receptors in regulation of respiration.

Spinal ventral root after-discharges as a pain index: Involvement of NK-1 and NMDA receptors

Nociceptive signals are transmitted to the spinal dorsal horn via primary afferent fibers, and the signals induce withdrawal reflexes by activating spinal motoneurons in the ventral horn. Therefore, nociceptive stimuli increase motoneuronal firing and ventral root discharges. This study was aimed to develop a method for the study of pain mechanisms and analgesics by recording ventral root discharges. Spinalized rats were laminectomized in the lumbo-sacral region. The fifth lumbar ventral root was sectioned and placed on a pair of wire electrodes. Multi unit efferent discharges from the ventral root were increased by mechanical stimulation using a von Frey hair applied to the plantar surface of the hindpaw. The low-intensity mechanical stimuli increased the discharges during stimulation (during-discharges) without increasing the discharges after cessation of stimulation (after-discharges), and the high-intensity mechanical stimuli increased both during- and after-discharges. Pretreatment with resiniferatoxin, an ultrapotent analogue of capsaicin, halved during-discharges and eliminated after-discharges, suggesting that after-discharges are generated by heat- and mechanosensitive polymodal nociceptors. Ezlopitant, a neurokinin-1 (NK-1) receptor antagonist, but not its inactive enantiomer, selectively reduced the after-discharges. Ketamine, an N-methyl- d-aspartate (NMDA) receptor antagonist, preferentially reduced the after-discharges, demonstrating that NK-1 and NMDA receptors mediate the after-discharges. Morphine reduced the after-discharges without affecting during-discharges. By contrast, mephenesin, a centrally acting muscle relaxant, reduced both during- and after-discharges. There results suggest that simultaneous recordings of during- and after-discharges are useful to study pain mechanisms and analgesics as well as to discriminate the analgesic effects from the side effects such as muscle relaxant effects.

Fictive locomotor patterns generated by tetraethylammonium application to the neonatal rat spinal cord in vitro

Intrinsic spinal networks generate a locomotor rhythm characterized by alternating electrical discharges from flexor and extensor motor pools. Because this process is preserved in the rat isolated spinal cord, this preparation in vitro may be a useful model to explore methods to reactivate locomotor networks damaged by spinal injury. The present electrophysiological investigation examined whether the broad spectrum potassium channel blocker tetraethylammonium could generate locomotor-like patterns. Low (50–500μM) concentrations of tetraethylammonium induced irregular, synchronous discharges incompatible with locomotion. Higher concentrations (1–10mM) evoked alternating discharges between flexor and extensor motor pools, plus large depolarization of motoneurons with spike broadening. The alternating discharges were superimposed on slow, shallow waves of synchronous depolarization. Rhythmic alternating patterns were suppressed by blockers of glutamate, GABA A and glycine receptors, disclosing a background of depolarizing bursts inhibited by antagonism of group I metabotropic glutamate receptors. Furthermore, tetraethylammonium also evoked irregular discharges on dorsal roots. Rhythmic alternating patterns elicited by tetraethylammonium on ventral roots were relatively stereotypic, had limited synergy with fictive locomotion induced by dorsal root stimuli, and were not accelerated by 4-aminopyridine. Horizontal section of the spinal cord preserved irregular ventral root discharges and dorsal root discharges, demonstrating that the action of tetraethylammonium on spinal networks was fundamentally different from that of 4-aminopyridine. These results show that a potassium channel blocker such as tetraethylammonium could activate fictive locomotion in the rat isolated spinal cord, although the pattern quality lacked certain features like frequency modulation and strong synergy with other inputs to locomotor networks.

Is NMDA Receptor Activation Essential for the Production of Locomotor-Like Activity in the Neonatal Rat Spinal Cord?

Previous work has established that in vitro bath application of N-methyl-D-aspartic acid (NMDA) promotes locomotor activity in a variety of vertebrate preparations including the neonatal rat spinal cord. In addition, NMDA receptor activation gives rise to active membrane properties that are postulated to contribute to the generation or stabilization of locomotor rhythm. However, earlier studies yielded conflicting evidence as to whether NMDA receptors are essential in this role. Therefore in this study, we examined the effect of NMDA receptor blockade, using D-2-amino-5-phosphono-valeric acid (AP5), on locomotor-like activity in the in vitro neonatal rat spinal cord. Locomotor-like activity was induced using 5-hydroxytryptamine (5-HT), acetylcholine, combined 5-HT and NMDA receptor activation, increased K(+) concentration, or electrical stimulation of the brain stem and monitored using suction electrode recordings of left and right lumbar ventral root discharge. We also studied the effect on locomotor capacity of selectively suppressing NMDA receptor-mediated active membrane properties; this was achieved by removing Mg(2+) ions from the bath, which in turn abolishes voltage-sensitive blockade of the NMDA receptor channel. The results show that, although NMDA receptor activation may seem essential for locomotor network operation under some experimental conditions, locomotor-like rhythms can nevertheless be generated in the presence of AP5 if spinal cord circuitry is exposed to appropriate levels of non-NMDA receptor-dependent excitation. Therefore neither NMDA receptor-mediated nonlinear membrane properties nor NMDA receptor activation in general is universally essential for locomotor network activation in the in vitro neonatal rat spinal cord.

Two Methods of Modeling of Spinal Superreflexia in Rats

We studied the relations between conditions of induction and quantitative characteristics of abnormally amplified monosynaptic reflex discharges (MSD) in ventral roots (VR) observed in two experimental situations: (i) 5 days after simultaneously performed denervation (transection of the sciatic nerve) and spinalization at the L1 level, and (ii) 5 days after preliminary denervation and with systemic injection of 4-aminopyridine (4-AP) in the course of the acute experiment. In both situations, the amplitude of the MSD conducted via the VR was close to the threshold of excitation of fibers in this root or even exceeded this value (a superreflexia phenomenon). Under both (i) and (ii) conditions, we observed generation of the second component of MSD in the VR, which was probably related to transition of excitation from excited to “silent” fibers in the VR. The latter of the above variants of induction superreflexia (5 days after denervation and with the effect of 4-AP in the acute experiment) is preferred because there is practically no death of the experimental animals in the course of the chronic experiment, there are no negative post-spinalization changes in the spinal cord, and the possibility of supraspinal activation of motoneurons is preserved.

Induction of a non-rhythmic motor pattern by nitric oxide in hatchling Rana temporaria embryos.

Nitric oxide (NO) is a ubiquitous neuromodulator with a diverse array of functions in a variety of brain regions, but a role for NO in the generation of locomotor activity has yet to be demonstrated. The possibility that NO is involved in the generation of motor activity in embryos of the frog Rana temporaria was investigated using the NO donors S-nitroso-n-acetylpenicillamine (SNAP; 100--500 micromol l(-1)) and diethylamine nitric oxide complex sodium (DEANO; 25--100 micromol l(-1)). Immobilised Rana temporaria embryos generate a non-rhythmic 'lashing' motor pattern either spontaneously or in response to dimming of the experimental bath illumination. Bath-applied NO donors triggered a qualitatively similar motor pattern in which non-rhythmic motor bursts were generated contra- and ipsilaterally down the length of the body. The inactive precursor of SNAP, n-acetyl-penicillamine (NAP), at equivalent concentrations did not trigger motor activity. NO donors failed to initiate swimming and had no measurable effects on the parameters of swimming induced by electrical stimulation. Intracellular recordings with potassium-acetate-filled electrodes revealed that the bursts of ventral root discharge induced by NO donors were accompanied by phasic depolarisations in motor neurons. During the inter-burst intervals, periods of substantial membrane hyperpolarization below the normal resting potential were observed, presumably coincident with contralateral ventral root activity. With KCl-filled electrodes, inhibitory potentials were strongly depolarising, suggesting that inhibition was Cl(-)-dependent. The synaptic drive seen in motor neurons after dimming of the illumination was very similar to that induced by the NO donors. NADPH-diaphorase histochemistry identified putative endogenous sources of NO in the central nervous system and the skin. Three populations of bilaterally symmetrical neurons were identified within the brainstem. Some of these neurons had contralateral projections and many had axonal processes that projected to and entered the marginal zones of the spinal cord, suggesting that they were reticulospinal.

Properties of rhythmic activity generated by the isolated spinal cord of the neonatal mouse.

We examined the ability of the isolated lumbosacral spinal cord of the neonatal mouse (P0-7) to generate rhythmic motor activity under several different conditions. In the absence of electrical or pharmacological stimulation, we recorded several patterns of spontaneous ventral root depolarization and discharge. Spontaneous, alternating discharge between contralateral ventral roots could occur two to three times over a 10-min interval. We also observed other patterns, including left-right synchrony and rhythmic activity restricted to one side of the cord. Trains of stimuli delivered to the lumbar/coccygeal dorsal roots or the sural nerve reliably evoked episodes of rhythmic activity. During these evoked episodes, rhythmic ventral root discharges could occur on one side of the cord or could alternate from side to side. Bath application of a combination of N-methyl-D,L-aspartate (NMA), serotonin, and dopamine produced rhythmic activity that could last for several hours. Under these conditions, the discharge recorded from the left and right L(1)-L(3) ventral roots alternated. In the L(4)-L(5) segments, the discharge had two peaks in each cycle, coincident with discharge of the ipsilateral and contralateral L(1)-L(3) roots. The L(6) ventral root discharge alternated with that recorded from the ipsilateral L(1)-L(3) roots. We established that the drug-induced rhythm was locomotor-like by recording an alternating pattern of discharge between ipsilateral flexor and extensor hindlimb muscle nerves. In addition, by recording simultaneously from ventral roots and muscle nerves, we established that ankle flexor discharge was in phase with ipsilateral L(1)/L(2) ventral root discharge, while extensor discharge was in phase with ipsilateral L(6) ventral root discharge. Rhythmic patterns of ventral root discharge were preserved following mid-sagittal section of the spinal cord, demonstrating that reciprocal inhibitory connections between the left and right sides of the cord are not essential for rhythmogenesis in the neonatal mouse cord. Blocking N-methyl-D-aspartate receptors, in both the intact and the hemisected preparation, revealed that these receptors contribute to but are not essential for rhythmogenesis. In contrast, the rhythm was abolished following blockade of kainate/AMPA receptors with 6-cyano-7-nitroquinoxalene-2,3-dione. These findings demonstrate that the isolated mouse spinal cord can produce a variety of coordinated activities, including locomotor-like activity. The ability to study these behaviors under a variety of different conditions offers promise for future studies of rhythmogenic mechanisms in this preparation.

Locomotor network maturation is transiently delayed in the MAOA-deficient mouse.

In vivo and in vitro experiments were performed in control (C3H) and monoamine oxidase A (MAOA)-deficient (Tg8) neonatal mice to determine whether MAOA deficiency affected spinal locomotor network maturation. Comparing the swimming behaviors at birth in C3H mice with those in Tg8 mice revealed a delayed role for the hindlimbs in Tg8 swimming, even though adult swimming behavior was acquired at postnatal day 14 (P14) in both strains. Analyzing the locomotor network activity in vitro showed that serotonin (5-HT) induced and modulated locomotor-like discharges in hindlimb ventral roots of C3H but not Tg8 neonates. The Tg8 network began, however, to be affected by 5-HT at P11. Thus both in vivo and in vitro results argue for a transient delay of locomotor network maturation in the Tg8 strain.

Whole cell recordings of lumbar motoneurons during locomotor-like activity in the in vitro neonatal rat spinal cord.

Whole cell current- and voltage-clamp recordings were obtained from lumbar motoneurons in the isolated neonatal rat spinal cord to characterize the behavior of motoneurons during neurochemically induced locomotor-like activity. Bath application of serotonin (10-100 muM) in combination with N-methyl-D-aspartate (1-12 muM) initially produced tonic membrane depolarization (mean = 26 mV), increased input resistance, decreased rheobase, and increased spike inactivation in response to depolarizing current pulse injections. After the initial tonic depolarization, rhythmic fluctuations of the motoneuron membrane potential (locomotor drive potentials; LDPs) developed that were modulated phasically in association with ventral root discharge. The peak and trough voltage levels of the LDP fluctuated above and below the membrane potential recorded immediately before the onset of rhythmic activity. Similarly, firing frequency was modulated above and below prelocomotion firing rates (in those motoneurons that displayed neurochemically induced tonic firing immediately before the onset of rhythmic activity). These observations are consistent with an alternation between phasic excitatory and inhibitory synaptic drives. The amplitude of LDPs and rhythmic excitatory drive current increased with membrane depolarization from -80 to -40 mV and then decreased with further depolarization, thus displaying nonlinear voltage-dependence. Faster frequency, small amplitude voltage fluctuations were observed superimposed on the depolarized phase of LDPs. In some motoneurons, the trajectory of these superimposed fluctuations was consistent with a synaptic origin, whereas in other cells, the regular sinusoidal appearance of the fluctuations and the occurrence of superimposed plateau potentials were more compatible with the activation of an intrinsic membrane property. One motoneuron displayed exclusively excitatory phasic drive, and another motoneuron was characterized by inhibitory phasic drive alone, during rhythmic activity. These findings are compatible with the concept of a central pattern generator that is capable of delivering both excitatory and inhibitory drive to motoneurons during locomotion. The data also suggest that the rhythmic excitatory and inhibitory outputs of the hypothetical half-center model can be dissociated and operate in isolation.

Spatiotemporal characteristics of 5-HT and dopamine-induced rhythmic hindlimb activity in the in vitro neonatal rat

1. Rhythmic activity was induced with either serotonin (5-HT; 10-100 microM) or dopamine (0.1-1.0 mM), in the in vitro spinal cord preparation of neonatal rats, with one intact hindlimb attached. Patterns of activity were investigated with multiple EMG recordings and the spatiotemporal characteristics of 5-HT- and dopamine-induced activity compared. 2. Dopamine-induced rhythmic activity was slow (cycle duration: 2.2-70.1 s) and irregular, whereas rhythmic activity induced by 5-HT was fast (cycle duration: 1.3-5.1 s) and regular. 3. During 5-HT- and dopamine-induced rhythmic activity, the timing of muscular activity was similar for hip flexors and hip adductors, for semimembranosus (hip extensor), and for muscles controlling the ankle and the foot. 4. In contrast, notable differences in the phase in the pattern induced by 5-HT compared with that induced by dopamine were found in the biceps femoris, semitendinosus, and quadriceps muscles. Biceps femoris and semitendinosus (functional hip extensors and knee flexors) were always extensor-like during 5-HT-induced activity, whereas in dopamine, these muscles displayed flexor-like bursts and double bursts as well as extensor-like bursts. Lack of EMG activity in biceps femoris and semitendinosus was encountered also in dopamine. In rectus femoris, vastus lateralis, and vastus medialis (main function: knee extension), the activity was dominated by flexor-like bursts in 5-HT, whereas in dopamine the activity was shifted to a predominantly extensor-like pattern. 5. The relationship between flexor and extensor burst duration and cycle duration was more variable than described for locomotor activity in adult animals. 6. The relative timing of muscle activity was stable from P0 to P4. The most important difference between rats aged 0-1 days and rats aged 2-4 days was a delayed flexor-extensor transition in older animals. 7. The complex timing of hindlimb muscle activity was relatively unchanged after transecting all dorsal roots. 8. Finally, the relationship between flexor and extensor activity and ventral root discharges was determined. It was found that the L2 ventral root burst was in phase with simple flexors while the L5 burst coincide with the extensor phase. 9. We conclude, that 5-HT and dopamine can activate spinal central pattern generators (CPGs) that already at birth are able to produce distinct patterns of motor activity. Modulatory inputs thus seems to be able to reconfigure the CPGs to produce specific motor outputs.

Neurite outgrowth through lesions of neonatal opossum spinal cord in culture.

The aim of these experiments was to analyze neurite outgrowth during regeneration of opossum spinal cord isolated from Monodelfis domestica and maintained in culture for 3-5 days. Lesions were made by crushing with forceps. In isolated spinal cords of animals aged 3 days, neurites entered the crush and grew along the basal lamina of the pia mater. Growth cones with pleiomorphic appearance containing vesicles, mitochondria and microtubules were abundant in the marginal zone, as were synaptoid contacts with active zones facing basal lamina. In preparations from animals aged 11-12 days, the lesion site was disrupted and contained only degenerating axons, debris and vesicles. Axons and growth cones entered the edge of the lesion but did not extend into it. Lesions in young animals extended over distances of more than 1 mm and contained no radial glia. The damaged area in older preparations was restricted to the crush site with normal astrocytes, oligodendrocytes and neurons immediately adjacent to the lesion. Thus, similar crushes produced more extensive damage in younger spinal cords that were capable of regeneration than in older cords that were not. Dorsal root ganglion fibers labeled with carbocyanine dye (DiI) were observed by video imaging as they grew through lesions. Individual growth cones examined subsequently by electron microscopy had grown again along pial basal lamina. After 5 days in culture dorsal root stimulation gave rise to discharges in ventral roots beyond the lesion indicating that synaptic connections were formed by growing fibers.

Effects of dexamethasone on the electrical activity of spinal neurons in rats with the transected sciatic nerve

Effects of the glucocorticoid hormone dexamethasone on the reflex discharges in the lumbar ventral roots and background activity (BA) of single neurons in the dorsal laminae of spinal grey were studied in rats after transection of the sciatic nerve. Administration of the hormone during early post-traumatic period (up to seven days) evoked no significant changes in the amplitude of increased (due to the postdenervation hyperreflexia) monosynaptic discharges on the side of nerve transection. At the same time, the monosynaptic discharges grew by 150–170% on the intact side. During later post-transection periods (up to 35 days), when ventral root reflex discharges were suppressed, dexamethasone facilitated reflex transmission via the polysynaptic segmental pathways on both the operated and intact sides. Nonetheless, the monosynaptic component of reflex discharges on the injured side did not recover. Dexamethasone treatment resulted in an increase in the number of BA-generating interneurons within the superficial dorsal horn laminae, and in a decrease in the proportion of units generating bursting activity (possibly of pathological nature).

Evidence for the existence of a functional polysynaptic pathway from trigeminal afferents to lumbar motoneurons in the neonatal rat.

Stimulation of trigeminal afferents has been reported to have powerful effects on the spinal cord in adult animals of several species. In the present study, the pathway transmitting these influences was investigated in the neonatal rat. Experiments were performed on in vitro brainstem/spinal cord preparations. Stimulation of the trigeminal nerve evoked bilateral polysynaptic discharges in lumbar ventral roots. Intracellular recordings from lumbar motoneurons showed mainly excitatory responses, although a few inhibitory responses were also observed. Experiments with perfusion of different parts of the preparation with general or selective synaptic blockers revealed a synaptic relay under GABAergic control in the brainstem, and at least one synapse in the cervical and in the thoracic spinal cord. The involvement of lumbar interneurons was established by perfusing the lumbar enlargement with saline containing either a high concentration of divalent ions or mephenesin in order to reduce transmission along polysynaptic pathways. The contribution of excitatory amino acid transmission was evaluated and was found to evoke mixed receptor responses. The course of the pathway was traced by using different lesions to the brainstem and spinal cord. The pathway was found to be ipsilateral in the brainstem and to become bilateral in the spinal cord. The results of the present study demonstrate that polysynaptic sensorimotor pathways are present at birth. The results are discussed in relation to the pontomedullary locomotor strip, which has been thought to share many features with the trigeminal system.

A role for potassium currents in the generation of the swimming motor pattern of Xenopus embryos.

1. To assess the role that K+ currents play in the production of the swimming motor pattern in the Xenopus embryo, we have used low doses of the K+ channel blockers, 3,4-diaminopyridine (3,4-DAP; 25-100 microM) and tetraethylammonium (TEA; 500 microM), to reduce K+ currents and investigated the effects on motor output. 2. To confirm that 3,4-DAP and TEA block K+ currents and characterize their actions, we made whole-cell voltage-clamp recordings from acutely isolated spinal neurons. Both 25-100 microM 3,4-DAP and 100-500 microM TEA blocked the sustained K+ current in a dose-dependent manner. 3. Because TEA can block acetylcholine nicotinic receptors on autonomic ganglia, and nicotinic acetylcholine receptors have recently been shown to be present on Xenopus spinal neurons, we have tested both 3,4-DAP and TEA for antagonist action against the nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium (DMPP). Although 500 microM TEA blocked the DMPP-induced depolarization, 25 microM 3,4-DAP did not. 4. In the intact embryo, application of 25-100 microM 3,4-DAP or 500 microM TEA disrupted both the left and right alternation of ventral root discharge and the motor pattern recorded intracellularly from spinal neurons during swimming. Both blockers allowed the firing of an extra action potential at midcycle, which led to a number of different patterns. These patterns were categorized as follows: type A, cycles with midcycle action potentials; type B, the simultaneous firing of neurons on both sides of the cord; and type C, in which one side was active, whereas the other side was inhibited. In both 3,4-DAP and TEA these abnormalities tended to occur at the beginning of swimming episodes. Both blockers also caused a significant increase in the cycle period. Because both 3,4-DAP and TEA produced very similar affects to the motor pattern, we conclude that the perturbations are probably a result of reducing K+ current amplitude. 5. To investigate whether 3,4-DAP and TEA were producing disruptions in the motor pattern by increasing synaptic drive through the broadening of action potentials, we made measurements of spike width, tonic depolarization, and midcycle inhibitory postsynaptic potential (IPSP) amplitude during swimming. Both 3,4-DAP and TEA caused significant but modest spike broadening (20.8 and 29.8%, respectively); however, their effects on tonic depolarization were inconsistent although both blockers increased midcycle IPSP amplitude. 6. To test whether a reduction in K+ currents could plausibly produce the specific motor pattern perturbations that were seen, we have made computer simulations of simplified spinal networks.(ABSTRACT TRUNCATED AT 400 WORDS)