Postnatal Changes in K+/Cl- Cotransporter-2 Expression in the Forebrain of Mice Bearing a Mutant Nicotinic Subunit Linked to Sleep-Related Epilepsy

ABSTRACT The Na+/K+/Cl- cotransporter-1 (NKCC1) and the K+/Cl- cotransporter-2 (KCC2) set the transmembrane Cl- gradient in the brain, and are implicated in epileptogenesis. We studied the postnatal distribution of NKCC1 and KCC2 in wild-type (WT) mice, and in a mouse model of sleep-related epilepsy, carrying the mutant \u3b22-V287L subunit of the nicotinic acetylcholine receptor (nAChR). In WT neocortex, immunohistochemistry showed a wide distribution of NKCC1 in neurons and astrocytes. At birth, KCC2 was localized in neuronal somata, whereas at subsequent stages it was mainly found in the somatodendritic compartment. The cotransporters\u2019 expression was quantified by densitometry in the transgenic strain. KCC2 expression increased during the first postnatal weeks, while the NKCC1 amount remained stable, after birth. In mice expressing \u3b22-V287L, the KCC2 amount in layer V of prefrontal cortex (PFC) was lower than in the control littermates at postnatal day 8 (P8), with no concomitant change in NKCC1. Consistently, the GABAergic excitatory to inhibitory switch was delayed in PFC layer V of mice carrying \u3b22-V287L. At P60, the amount of KCC2 was instead higher in mice bearing the transgene. Irrespective of genotype, NKCC1 and KCC2 were abundantly expressed in the neuropil of most thalamic nuclei since birth. However, KCC2 expression decreased by P60 in the reticular nucleus, and more so in mice expressing \u3b22-V287L. Therefore, a complex regulatory interplay occurs between heteromeric nAChRs and KCC2 in postnatal forebrain. The pathogenetic effect of \u3b22-V287L may depend on altered KCC2 amounts in PFC during synaptogenesis, as well as in mature thalamocortical circuits

Determinants of synaptic integration and heterogeneity in rebound firing explored with data-driven models of deep cerebellar nucleus cells

Significant inroads have been made to understand cerebellar cortical processing but neural coding at the output stage of the cerebellum in the deep cerebellar nuclei (DCN) remains poorly understood. The DCN are unlikely to just present a relay nucleus because Purkinje cell inhibition has to be turned into an excitatory output signal, and DCN neurons exhibit complex intrinsic properties. In particular, DCN neurons exhibit a range of rebound spiking properties following hyperpolarizing current injection, raising the question how this could contribute to signal processing in behaving animals. Computer modeling presents an ideal tool to investigate how intrinsic voltage-gated conductances in DCN neurons could generate the heterogeneous firing behavior observed, and what input conditions could result in rebound responses. To enable such an investigation we built a compartmental DCN neuron model with a full dendritic morphology and appropriate active conductances. We generated a good match of our simulations with DCN current clamp data we recorded in acute slices, including the heterogeneity in the rebound responses. We then examined how inhibitory and excitatory synaptic input interacted with these intrinsic conductances to control DCN firing. We found that the output spiking of the model reflected the ongoing balance of excitatory and inhibitory input rates and that changing the level of inhibition performed an additive operation. Rebound firing following strong Purkinje cell input bursts was also possible, but only if the chloride reversal potential was more negative than −70 mV to allow de-inactivation of rebound currents. Fast rebound bursts due to T-type calcium current and slow rebounds due to persistent sodium current could be differentially regulated by synaptic input, and the pattern of these rebounds was further influenced by HCN current. Our findings suggest that active properties of DCN neurons could play a crucial role for signal processing in the cerebellum

Modulation of glutamate release by nicotinic receptors in layer V of the murine prefrontal cortex

By regulating the neocortical excitability, nicotinic acetylcholine receptors (nAChRs) control vigilance and cognition. In rodents, the neocortex mainly expresses homomeric a7 and heteromeric a4 f2 nAChRs. These are expressed in both pre- and postsynaptic locations and mediate classical synaptic transmission as well as slower paracrine effects. We have studied the contribution of heteromeric nAChRs to the control of glutamate (GLU) release in layer V of the murine prefrontal cortex (PFC). Tonic application of 5 \ub5M nicotine more than doubled the frequency of spontaneous glutamatergic excitatory postsynaptic currents recorded on pyramidal neurons in acute brain slices. The effect of nicotine was inhibited by 1 \ub5M dihydro- f-erythroidine (DH fE, which blocks a4-containing receptors), but not by 10 nM methyllicaconitine (MLA, which blocks a7-containing receptors). We next studied the association of a4 with different populations of glutamatergic terminals, in both PFC and somatosensory cortex. The GLU transporter type 1 (VGLUT1) mostly labels the intrinsic glutamatergic terminals or cortical afferents, whereas the type 2 transporter VGLUT2 tends to label the thalamic afferents. Immunofluorescence showed that a4 was expressed in both VGLUT1 and VGLUT2 terminals and colocalization was considerably stronger in the PFC. Expression of the a4, VGLUT1 and VGLUT2 was also tested by immunoblots, which confirmed the overall higher expression of these proteins in the PFC compared to the somatosensory cortex. Hence, in PFC, a4-containing heteromeric nAChRs are expressed in both intrinsic and extrinsic glutamatergic terminals and regulate GLU release in the presence of steady agonist levels