Daniel Cattaert

Down-regulation of the potassium-chloride cotransporter KCC2 contributes to spasticity after spinal cord injury

Hyperexcitability of spinal reflexes and reduced synaptic inhibition are commonly associated with spasticity after spinal cord injury (SCI). In adults, the activation of γ-aminobutyric acidA (GABAA) and glycine receptors inhibits neurons as a result of low intracellular chloride (Cl−) concentration, which is maintained by the potassium-chloride cotransporter KCC2 (encoded by Slc12a5). We show that KCC2 is downregulated after SCI in rats, particularly in motoneuron membranes, thereby depolarizing the Cl− equilibrium potential and reducing the strength of postsynaptic inhibition. Blocking KCC2 in intact rats reduces the rate-dependent depression (RDD) of the Hoffmann reflex, as is observed in spasticity. RDD is also decreased in KCC2-deficient mice and in intact rats after intrathecal brain-derived neurotrophic factor (BDNF) injection, which downregulates KCC2. The early decrease in KCC2 after SCI is prevented by sequestering BDNF at the time of SCI. Conversely, after SCI, BDNF upregulates KCC2 and restores RDD. Our results open new perspectives for the development of therapeutic strategies to alleviate spasticity.

Central control components of a 'simple' stretch reflex.

The monosynaptic stretch reflex is a fundamental feature of sensory-motor organization in most animal groups. In isolation, it serves largely as a negative feedback devoted to postural controls; however, when it is involved in diverse movements, it can be modified by central command circuits. In order to understand the implications of such modifications, a model system has been chosen that has been studied at many different levels: the crayfish walking system. Recent studies have revealed several levels of control and modulation (for example, at the levels of the sensory afferent and the output synapse from the sensory afferent, and via changes in the membrane properties of the postsynaptic neuron) that operate complex and highly adaptive sensory-motor processing. During a given motor task, such mechanisms reshape the sensory message completely, such that the stretch reflex becomes a part of the central motor command.

Monosynaptic Interjoint Reflexes and their Central Modulation During Fictive Locomotion in Crayfish.

An in vitro preparation of the crayfish nervous system has been utilized to study an interjoint reflex pathway and its variability during rhythmic locomotor activity. The coxo‐basal chordotonal organ (CBCO) is a joint stretch receptor spanning the second joint of walking legs in crayfish, where it encodes joint movements and position. Mechanical stimulation (stretch and release) of the CBCO and electrical stimulation of the CBCO nerve elicits reflex responses in promotor and remotor motor neurons innervating muscles moving the basal thoraco‐coxal (TC) leg joint. Promotor and remotor motor neurons receive monosynaptic excitatory inputs from at least four CBCO afferents, including both stretch‐ and release‐sensitive CBCO afferents. In a tonic preparation, in which there is no tendency to produce alternating bursts of activity in antagonistic motor neurons, the reflex responses were evoked during each cycle of imposed movement. However, when the preparation became rhythmic and produced bouts of fictive locomotion, the reflex responses were unstable and their gain was phasically modulated. Paired recordings indicate that such a modulation of the monosynaptic interjoint reflex could be due to both a phasic change in the excitability of the motor neurons and presynaptic inhibition that reduces the excitatory input from CBCO primary afferents.

Simulating a neural cross-talk model for between-hand interference during bimanual circle drawing.

Abstract. Studies on drawing circles with both hands in the horizontal plane have shown that this task is easy to perform across a wide range of movement frequencies under the symmetrical mode of coordination, whereas under the asymmetrical mode (both limbs moving clockwise or counterclockwise) increases in movement frequency have a disruptive effect on trajectory control and hand coordination. To account for these interference effects, we propose a simplified computer model for bimanual circle drawing based on the assumptions that (1) circular trajectories are generated from two orthogonal oscillations coupled with a phase delay, (2) the trajectories are organized on two levels, “intention” and “motor execution”, and (3) the motor systems controlling each hand are prone to neural cross-talk. The neural cross-talk consists in dispatching some fraction of any force command sent to one limb as a mirror image to the other limb. Assuming predominating coupling influences from the dominant to the nondominant limb, the simulations successfully reproduced the main characteristics of performance during asymmetrical bimanual circle drawing with increasing movement frequencies, including disruption of the circular form drawn with the nondominant hand, increasing dephasing of the hand movements, increasing variability of the phase difference, and occasional reversals of the movement direction in the nondominant limb. The implications of these results for current theories of bimanual coordination are discussed.

Dual personality of GABA/glycine-mediated depolarizations in immature spinal cord

The inhibitory action of glycine and GABA in adult neurons consists of both shunting incoming excitations and moving the membrane potential away from the action potential (AP) threshold. By contrast, in immature neurons, inhibitory postsynaptic potentials (IPSPs) are depolarizing; it is generally accepted that, despite their depolarizing action, these IPSPs are inhibitory because of the shunting action of the Cl− conductance increase. Here we investigated the integration of depolarizing IPSPs (dIPSPs) with excitatory inputs in the neonatal rodent spinal cord by means of both intracellular recordings from lumbar motoneurons and a simulation using the compartment model program “Neuron.” We show that the ability of IPSPs to suppress suprathreshold excitatory events depends on ECl and the location of inhibitory synapses. The depolarization outlasts the conductance changes and spreads electrotonically in the somatodendritic tree, whereas the shunting effect is restricted and local. As a consequence, dIPSPs facilitated AP generation by subthreshold excitatory events in the late phase of the response. The window of facilitation became wider as ECl was more depolarized and started earlier as inhibitory synapses were moved away from the excitatory input. GAD65/67 immunohistochemistry demonstrated the existence of distal inhibitory synapses on motoneurons in the neonatal rodent spinal cord. This study demonstrates that small dIPSPs can either inhibit or facilitate excitatory inputs depending on timing and location. Our results raise the possibility that inhibitory synapses exert a facilitatory action on distant excitatory inputs and slight changes of ECl may have important consequences for network processing.

Monosynaptic connections mediate resistance reflex in crayfish (Procambarus clarkii) walking legs

SummaryIn crustacean walking legs, the coxo-basipodite chordotonal organ (CB) composed of about 50 sensory cells, evokes a resistance reflex in the levator (Lev) and depressor (Dep) muscles responsible for the movements of the coxo-basipodite joint where it is located. Mechanical stimulation of the CB strand and electrical stimulation of its sensory nerve have been performed along with systematic intracellular recordings from CB terminals (CB T) and levator (Lev) or depressor (Dep) motoneurons (MNs) in order to study their connections. Measurements of conduction times in the CB nerve demonstrated different pools of sensory fibres, the fastest of which reach the ganglion in 2.5 ms. During imposed movements to the CB strand, intracellularly recorded Lev or Dep MN display EPSPs that are correlated to spikes in the CB nerve, their delays are incompatible with a polysynaptic pathway. Systematic stimulation of the CB nerve demonstrates that about 4 to 8 CB fibres are connected with each Lev or Dep MN. Classical tests for monosynaptic connections indicate that EPSPs occurring between 3 ms and 6 ms correspond mainly to monosynaptic connections with CB T, whereas IPSPs (the latencies of which are above 12 ms) are polysynaptic. In spite of the high selectivity of the CB T onto MNs, eight simultaneous intracellular recordings of coupled CB T and MN (out of more than 300 MNs penetrated) have allowed a direct measurement of synaptic delays (less than 1 ms). The functional significance of these results is discussed in relation to the proprioceptive control of locomotor movements.

Anxiety-like behavior in crayfish is controlled by serotonin

The crayfish that was afraid of the dark We tend to assume that complex emotions, such as anxiety, only occur in mammals or other cognitively complex vertebrates. But a heightened sense of awareness and the avoidance of novel or dangerous environments could be helpful for any animal species. Fossat et al. show that crayfish exposed to a stressful electric field refuse to enter dark arms in a light/dark maze, even after the electric field has been removed. The animals calmed down when they were injected with an anxiolytic drug used to treat anxiety in humans, and they entered the dark as normal. The stressed animals had increased levels of the neurotransmitter serotonin in the brain, and injections of serotonin induced anxiety-like behavior in control animals. Thus, these invertebrates display a primitive form of anxiety that shares a mechanism with the more complex emotions displayed by vertebrates. Science, this issue p. 1293 Crayfish respond to stress with an apparent fear of the dark that can be abolished with an anxiolytic drug. Anxiety, a behavioral consequence of stress, has been characterized in humans and some vertebrates, but not invertebrates. Here, we demonstrate that after exposure to stress, crayfish sustainably avoided the aversive illuminated arms of an aquatic plus-maze. This behavior was correlated with an increase in brain serotonin and was abolished by the injection of the benzodiazepine anxiolytic chlordiazepoxide. Serotonin injection into unstressed crayfish induced avoidance; again, this effect was reversed by injection with chlordiazepoxide. Our results demonstrate that crayfish exhibit a form of anxiety similar to that described in vertebrates, suggesting the conservation of several underlying mechanisms during evolution. Analyses of this ancestral behavior in a simple model reveal a new route to understanding anxiety and may alter our conceptions of the emotional status of invertebrates.

Chloride conductance produces both presynaptic inhibition and antidromic spikes in primary afferents

Primary afferents from a crayfish leg proprioceptor display both primary afferent depolarizations (PADs) and antidromic spikes. PADs are generated by activation of GABA receptors and produce presynaptic inhibition, while the antidromic spikes do not elicit any synaptic effect in the postsynaptic neurons. The aim of the present study was to investigate the ionic mechanisms that allow PADs to produce antidromic spikes and to test whether GABA can produce similar effects. Intracellular recordings from the sensory axon terminals within the ganglion where PAD are produced were performed. Lowering the extracellular chloride concentration resulted in an increase in PAD amplitude, which was then capable of producing antidromic spikes. Local application of GABA close to the axon terminal also resulted in production of antidromic spikes. We conclude that antidromic spikes may result from the activation of a GABA-mediated increase in chloride conductance that also produces PADs. Therefore PADs and antidromic spikes may represent two aspects of the same GABAergic inhibitory mechanism that gate sensory transmission.

Activity-dependent mismatch between axo-axonic synapses and the axon initial segment controls neuronal output

Significance Neurons integrate synaptic inputs at the axon initial segment (AIS), where the action potential is initiated. Recent findings have shown this structure to be highly plastic. Here, we focus on the axo-axonic synapses formed onto the AIS and show that although chronic increases in neuronal activity result in a distal relocation of the AIS, the synapses do not change position. Computational modeling suggests this spatial mismatch has critical functional consequences on neuronal output, allowing the cell to downregulate its own excitability and thus respond homeostatically to chronic stimulation. The axon initial segment (AIS) is a structure at the start of the axon with a high density of sodium and potassium channels that defines the site of action potential generation. It has recently been shown that this structure is plastic and can change its position along the axon, as well as its length, in a homeostatic manner. Chronic activity-deprivation paradigms in a chick auditory nucleus lead to a lengthening of the AIS and an increase in neuronal excitability. On the other hand, a long-term increase in activity in dissociated rat hippocampal neurons results in an outward movement of the AIS and a decrease in the cell’s excitability. Here, we investigated whether the AIS is capable of undergoing structural plasticity in rat hippocampal organotypic slices, which retain the diversity of neuronal cell types present at postnatal ages, including chandelier cells. These interneurons exclusively target the AIS of pyramidal neurons and form rows of presynaptic boutons along them. Stimulating individual CA1 pyramidal neurons that express channelrhodopsin-2 for 48 h leads to an outward shift of the AIS. Intriguingly, both the pre- and postsynaptic components of the axo-axonic synapses did not change position after AIS relocation. We used computational modeling to explore the functional consequences of this partial mismatch and found that it allows the GABAergic synapses to strongly oppose action potential generation, and thus downregulate pyramidal cell excitability. We propose that this spatial arrangement is the optimal configuration for a homeostatic response to long-term stimulation.

Embryonic alteration of motoneuronal morphology induces hyperexcitability in the mouse model of amyotrophic lateral sclerosis.

Although amyotrophic lateral sclerosis (ALS) is an age-dependent fatal neurodegenerative disease in which upper and lower motoneurons (MNs) are targeted for death in adults, increasing lines of evidence indicate that MNs display physiological and morphological abnormalities during postnatal development, long before disease onset. Here, using transgenic mice overexpressing the G93A mutation of the human Cu/Zn superoxide dismutase gene (SOD1), we show that SOD1(G93A) embryonic lumbar E17.5 MNs already expressed abnormal morphometric parameters, including a deep reduction of their terminal segments length. Whole-cell patch-clamp recordings from acute spinal cord preparations were made to characterize functional changes in neuronal activity. SOD1(G93A) E17.5 MNs displayed hyperexcitability compared to wild-type MNs. Finally, we performed realistic simulations in order to correlate morphometric and electrophysiological changes observed in embryonic SOD1(G93A) MNs. We found that the reduced dendritic elongation mainly accounted for the hyperexcitability observed in SOD1(G93A) MNs. Altogether, our results emphasize the remarkable early onset of abnormal neural activity in the commonly used animal model for ALS, and suggest that embryonic morphological changes are the primary compensatory mechanisms, the physiological adjustments being only secondary to morphological alterations.