Fictive Locomotion In Lamprey Research Articles

Intracellular QX-314 causes depression of membrane potential oscillations in lamprey spinal neurons during fictive locomotion.

Spinal neurons undergo large cyclic membrane potential oscillations during fictive locomotion in lamprey. It was investigated whether these oscillations were due only to synaptically driven excitatory and inhibitory potentials or if voltage-dependent inward conductances also contribute to the depolarizing phase by using N-(2,6-dimethylphenyl carbamoylmethyl)triethylammonium bromide (QX-314) administered intracellularly during fictive locomotion. QX-314 intracellularly blocks inactivating and persistent Na+ channels, and in some neurons, effects on certain other types of channels have been reported. To detail the effects of QX-314 on Na+ and Ca2+ channels, we used dissociated lamprey neurons recorded under whole cell voltage clamp. At low intracellular concentrations of QX-314 (0.2 mM), inactivating Na+ channels were blocked and no effects were exerted on Ca2+ channels (also at 0.5 mM). At 10 mM QX-314, there was, however a marked reduction of I(Ca). In the isolated spinal cord of the lamprey, fictive locomotion was induced by superfusing the spinal cord with Ringer's solution containing N-methyl-D-aspartate (NMDA), while recording the locomotor activity from the ventral roots. Simultaneously, identified spinal neurons were recorded intracellularly, while infusing QX-314 from the microelectrode. Patch electrodes cannot be used in the intact spinal cord, and therefore "sharp" electrodes were used. The amplitude of the oscillations was consistently reduced by 20-25% in motoneurons (P < 0.05) and unidentified spinal neurons (P < 0.005). The onset of the effect started a few minutes after impalement and reached a stable level within 30 min. These effects thus show that QX-314 causes a reduction in the amplitude of membrane potential oscillations during fictive locomotion. We also investigated whether QX-314 could affect glutamate currents by applying short pulses of glutamate from an extracellular pipette. No changes were observed. We also found no evidence for a persistent Na+ current in dissociated neurons, but these cells have a much-reduced dendritic tree. The results indicate that there is an inward conductance, which is sensitive to QX-314, during membrane potential oscillations that "boosts" the synaptic drive during fictive locomotion. Taken together, the results suggest that inactivating Na+ channels contribute to this inward conductance although persistent Na+ channels, if present on dendrites, could possibly also contribute to shaping the membrane potential oscillations.

The spinal 5-HT system contributes to the generation of fictive locomotion in lamprey

Activation of NMDA receptors evokes sustained fictive locomotion in the isolated spinal cord of the sea lamprey Petromyzon marinus ( P. marinus), but in the river lamprey Lampetra fluviatilis ( L. fluviatilis) the ventral root activity is often irregular. A previous study showed that the number of 5-HT immunoreactive fibres, neurones and varicosities are much lower in the spinal cord of L. fluviatilis than in P. marinus. To further analyse the underlying mechanisms, the present study investigated the role of the 5-HT system in stabilising fictive locomotion. In P. marinus a blockade of 5-HT1A receptors by spiperone reversibly increased the frequency and the coefficient of variation. This implies that there is an endogenous release of 5-HT during fictive locomotion that is important for the generation of locomotor activity. In L. fluviatilis bath applied NMDA or d-glutamate evoked in most cases irregular activity. An addition of 5-HT (0.5–2 μM) rapidly stabilised the burst generation and led to a sustained fictive locomotion. In a split-bath configuration, NMDA administered to the rostral part of the spinal cord in P. marinus evoked fictive locomotion in both the rostral part and the first few segments of the caudal part. When spiperone was added to the caudal part, the burst activity changed into tonic activity within 10 min. Taken together, these results indicate that activity in the intrinsic 5-HT system in the lamprey spinal locomotor network contributes significantly to the rhythm generation. The quantitative differences with regard to the 5-HT plexus between P. marinus and L. fluviatilis may account for the observed discrepancy between the two species.

Central modulation of stretch receptor neurons during fictive locomotion in lamprey.

1. In lamprey, stretch receptor neurons (SRNs), also referred to as edge cells, are located along the lateral margin of the spinal cord. They sense the lateral movements occurring in each swim cycle during locomotion. The isolated lamprey spinal cord in vitro was used to investigate the activity of SRNs during fictive locomotion induced by bath-applied N-methyl-D-aspartate (NMDA). Intracellular recordings with potassium acetate filled electrodes showed that 63% of SRNs had a clear locomotor-related modulation of their membrane potential. 2. Of the modulated SRNs, two-thirds had periods of alternating excitation and inhibition occurring during the ipsilateral and the contralateral ventral root bursts, respectively. The phasic hyperpolarization could be reversed into a depolarizing phase after the injection of chloride ions into the cells; this revealed a chloride-dependent synaptic drive. The remaining modulated SRNs were inhibited phasically during ipsilateral motor activity. 3. Experiments with barriers partitioning the recording chamber with the spinal cord into three pools, allowed an inactivation of the locomotor networks within one pool by washing out NMDA from the pool in which the SRN was recorded. This resulted in a marked reduction, but not an abolishment, of the amplitude of the membrane potential oscillations. Both the excitatory and the inhibitory phases were reduced, resulting from removal of input from inhibitory and excitatory interneurons projecting from the adjacent pools. If the glycine receptor antagonist strychnine (1 microM) was applied in one pool, the phasic hyperpolarizing phase disappeared without affecting the excitatory phase. 4. Bath application of the gamma-aminobutyric acid (GABA)A receptor antagonist, bicuculline (50-100 microM) blocked the spontaneous large unitary inhibitory postsynaptic potentials, which occurred without a clear phasic pattern. Bicuculline had no significant effect on the peak to peak amplitude of the locomotor-related membrane potential oscillations. The inhibition in SRNs therefore has a dual origin: glycinergic interneurons provide phasic inhibition, while the GABA system can exert a tonic inhibition via GABAA receptors. 5. These data show that, in addition to the stretch-evoked excitation, which SRNs receive during each locomotor cycle, most of them also receive excitation from the central pattern generator network during the ipsilateral contraction, which may ensure a maintained high level of sensitivity to stretch during the shortening phase of the locomotor cycle. This arrangement is analogous to the efferent control of muscle spindles exerted by gamma-motoneurons in mammals, which as a rule are coactivated with alpha-motoneurons to the same muscle (alpha-gamma linkage).

Modulation of swimming rhythmicity by 5-hydroxytryptamine during post-embryonic development in Xenopus laevis.

During the first 24 h of post-embryonic development in Xenopus laevis, a rapid change in the neural activity underlying swimming occurs in which the duration of ventral root discharge on each cycle increases from a single compound impulse to discrete bursts of activity. Moreover, this change in motor output progresses rostrocaudally, suggesting that it could result from the influence of a descending neural pathway upon the spinal rhythm-generating circuitry during early post-embryonic development. To begin to examine whether serotonergic neurons of brainstem raphe nuclei might have a role in this swimming development, we have studied the effects of 5-hydroxytryptamine (5HT) on fictive swimming in embryonic and larval animals. As previously demonstrated for other vertebrate locomotor rhythms, we find that bath-applied 5HT enhances the duration of motor activity on each cycle of larval fictive swimming. In addition, our results show that the sensitivity of the swimming rhythm to exogenous 5HT follows a strict rostrocaudal gradient. In young embryos (stages 32-36) 5HT does not affect the duration of ventral root impulses per cycle; by the time of hatching (stage 37/38), rostral but not caudal discharge is enhanced, and by stage 42 (24 h post-hatching) 5HT can increase motor burst durations along most of the length of the animal. These reversible changes induced by bath-applied 5HT closely resemble the normal rostrocaudal development of burst discharge during swimming in animals some 12 h older.(ABSTRACT TRUNCATED AT 250 WORDS)

Putative GABAergic input to axons of spinal interneurons and primary sensory neurons in the lamprey soinal cord as shown by intracellular Lucifer yellow and GABA immunohistochemistry

GABAergic plastic modulation of the membrane potential occurs in spinal interneurons during fictive locomotion in lamprey presumably indicating a presynaptic inhibition. GABA also modulates synaptic transmission from primary sensory neurons (dorsal cells) at a presynaptic site. From these findings GABA terminals would be expected to be in close contact with phasically modulated axons of spinal interneurons and/or dorsal cells and their axons. To test this supposition intracellular injections of Lucifer yellow into spinal interneurons or dorsal cells were combined with GABA immunohistochemistry. GABA-immunoreactive (ir) varicosities were found to be in close contact (<1 μm distance) with axons modulated during fictive locomotion as well as dorsal cell axons. Small GABA-ir bipolar neurons form processes, which are in close contact with the axons of dorsal cells.

Dorsal and ventral myotome motoneurons and their input during fictive locomotion in lamprey

Motoneurons supplying the dorsal and ventral parts of the myotome in the lamprey are shown to have different morphological characteristics; furthermore, their pattern of activation during fictive locomotion may differ considerably. Intracellular recordings from motoneurons were performed in an in vitro spinal cord-myotome preparation from segments rostral to the fins. The location of the contracting muscle fibers in the myotome could be observed directly in the dissection microscope during intracellular stimulation of the motoneuron. The motoneurons were injected with Lucifer Yellow, an intracellular dye, and were subsequently reconstructed, sometimes in both a horizontal and a transverse plane. Motoneurons supplying the ventral third of the myotome had a dense, fan-like, dendritic tree and ramifications near the midline. In contrast, motoneurons supplying the dorsal third of the myotome had a more widespread and less dense dendritic tree, with few ramifications near the midline. Some motoneurons supplying the most ventral or dorsal part of the myotome had contralateral dendrites crossing in the ventral commissure and ramifying near contralateral large, reticulospinal Müller fibers. The differences in morphology may indicate that these motoneurons receive different descending inputs. This may be related to the need for an effective control in the dorsoventral plane during righting and steering responses. During fictive locomotion elicited in the isolated spinal cord by bath-applied N-methyl-aspartate, pairs of motoneurons were recorded which subsequently were identified and characterized by intracellular injections of Lucifer Yellow.(ABSTRACT TRUNCATED AT 250 WORDS)