Hyperexcitability of spinal reflexes and reduced synaptic inhibition are commonly associated with spasticity after spinal cord injury (SCI). In adults, the activation of gamma-aminobutyric acid(A) (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.

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 E Cl 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 E Cl 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 E Cl may have important consequences for network processing.chloride homeostasis ͉ facilitation ͉ inhibition ͉ synaptic integration G ABA and glycine are excitatory in the immature spinal cord and become inhibitory during development. The shift from depolarizing to hyperpolarizing inhibitory postsynaptic potentials (IPSPs) occurs during the first postnatal week (1), a time window during which motoneurons (MNs) undergo considerable maturation of membrane properties (see ref. 2 for review). Some 15 years after the demonstration that GABA and glycine depolarize immature neurons (3-12), the excitatory or inhibitory nature of these depolarizations is still a matter of debate. A critical factor appears to be the equilibrium potential for Cl Ϫ ions (E Cl ) relative to action potential (AP) threshold. Spinal cord neurons in 4-to 7-day-old cultures exhibit spontaneous firing that is depressed by application of bicuculline to block GABA A receptors, suggesting that GABA release from developing axons can drive sodium APs (13). Similarly, a brief application of glycine onto the in vitro spinal cord isolated from fetal rats, at embryonic day 15.5 (i.e., 1 week before birth), evokes excitatory responses that are abolished by strychnine (14). Therefore, there is no doubt that GABA and glycine can play an excitatory role at an early stage of the development of spinal MNs and interneuron...

GABA and glycine are excitatory in the immature spinal cord and become inhibitory during development. The shift from depolarizing to hyperpolarizing inhibitory postsynaptic potentials (IPSPs) occurs during the perinatal period in the rat, a time window during which the projections from the brain stem reach the lumbar enlargement. In this study, we investigated the effects of suppressing influences of the brain on lumbar motoneurons during this critical period for the negative shift of the reversal potential of IPSPs (E(IPSP)). The spinal cord was transected at the thoracic level on the day of birth [postnatal day 0 (P0)]. E(IPSP), at P4-P7, was significantly more depolarized in cord-transected than in cord-intact animals (E(IPSP) above and below resting potential, respectively). E(IPSP) at P4-P7 in cord-transected animals was close to E(IPSP) at P0-P2. K-Cl cotransporter KCC2 immunohistochemistry revealed a developmental increase of staining in the area of lumbar motoneurons between P0 and P7 in cord-intact animals; this increase was not observed after spinal cord transection. The motoneurons recorded from cord-transected animals were less sensitive to the experimental manipulations aimed at testing the functionality of the KCC2 system, which is sensitive to [K(+)](o) and blocked by bumetanide. Although bumetanide significantly depolarized E(IPSP), the shift was less pronounced than in cord-intact animals. In addition, a reduction of [K(+)](o) affected E(IPSP) significantly only in cord-intact animals. Therefore influences from the brain stem may play an essential role in the maturation of inhibitory synaptic transmission, possibly by upregulating KCC2 and its functionality.

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