Forebrain-independent generation of hyperthermic convulsions in infant rats.

Febrile seizures are the most common type of convulsive events in children. It is generally assumed that the generalization of these seizures is a result of brainstem invasion by the initial limbic seizure activity. Using precollicular transection in 13-day-old rats to isolate the forebrain from the brainstem, we demonstrate that the forebrain is not required for generation of tonic-clonic convulsions induced by hyperthermia or kainate. Compared with sham-operated littermate controls, latency to onset of convulsions in both models was significantly shorter in pups that had undergone precollicular transection, indicating suppression of the brainstem seizure network by the forebrain in the intact animal. We have shown previously that febrile seizures are precipitated by hyperthermia-induced respiratory alkalosis. Here, we show that triggering of hyperthermia-induced hyperventilation and consequent convulsions in transected animals are blocked by diazepam. The present data suggest that the role of endogenous brainstem activity in triggering tonic-clonic seizures should be re-evaluated in standard experimental models of limbic seizures. Our work sheds new light on the mechanisms that generate febrile seizures in children and, therefore, on how they might be treated.

Neuronal carbonic anhydrase VII provides GABAergic excitatory drive to exacerbate febrile seizures.

Brain carbonic anhydrases (CAs) are known to modulate neuronal signalling. Using a novel CA VII (Car7) knockout (KO) mouse as well as a CA II (Car2) KO and a CA II/VII double KO, we show that mature hippocampal pyramidal neurons are endowed with two cytosolic isoforms. CA VII is predominantly expressed by neurons starting around postnatal day 10 (P10). The ubiquitous isoform II is expressed in neurons at P20. Both isoforms enhance bicarbonate-driven GABAergic excitation during intense GABAA-receptor activation. P13-14 CA VII KO mice show behavioural manifestations atypical of experimental febrile seizures (eFS) and a complete absence of electrographic seizures. A low dose of diazepam promotes eFS in P13-P14 rat pups, whereas seizures are blocked at higher concentrations that suppress breathing. Thus, the respiratory alkalosis-dependent eFS are exacerbated by GABAergic excitation. We found that CA VII mRNA is expressed in the human cerebral cortex before the age when febrile seizures (FS) occur in children. Our data indicate that CA VII is a key molecule in age-dependent neuronal pH regulation with consequent effects on generation of FS.

Subplate neurons promote spindle bursts and thalamocortical patterning in the neonatal rat somatosensory cortex.

Patterned neuronal activity such as spindle bursts in the neonatal cortex is likely to promote the maturation of cortical synapses and neuronal circuits. Previous work on cats has shown that removal of subplate neurons, a transient neuronal population in the immature cortex, prevents the functional maturation of thalamocortical and intracortical connectivity. Here we studied the effect of subplate removal in the neonatal rat primary somatosensory cortex (S1). Using intracortical EEG we show that after selective removal of subplate neurons in the limb region of S1, endogenous and sensory evoked spindle burst activity is largely abolished. Consistent with the reduced in vivo activity in the S1 limb region, we find by in vitro recordings that thalamocortical inputs to layer 4 neurons are weak. In addition, we find that removal of subplate neurons in the S1 barrel region prevents the development of the characteristic histological barrel-like appearance. Thus, subplate neurons are crucially involved in the generation of particular types of early network activity in the neonatal cortex, which are an important feature of cortical development. The altered EEG pattern following subplate damage could be applicable in the neurological assessment of human neonates.

Gene delivery to postnatal rat brain by non-ventricular plasmid injection and electroporation.

Creation of transgenic animals is a standard approach in studying functions of a gene of interest in vivo. However, many knockout or transgenic animals are not viable in those cases where the modified gene is expressed or deleted in the whole organism. Moreover, a variety of compensatory mechanisms often make it difficult to interpret the results. The compensatory effects can be alleviated by either timing the gene expression or limiting the amount of transfected cells. The method of postnatal non-ventricular microinjection and in vivo electroporation allows targeted delivery of genes, siRNA or dye molecules directly to a small region of interest in the newborn rodent brain. In contrast to conventional ventricular injection technique, this method allows transfection of non-migratory cell types. Animals transfected by means of the method described here can be used, for example, for two-photon in vivo imaging or in electrophysiological experiments on acute brain slices.