Cortical neurons and networks are dormant but fully responsive during isoelectric brain state.

A continuous isoelectric electroencephalogram reflects an interruption of endogenously-generated activity in cortical networks and systematically results in a complete dissolution of conscious processes. This electro-cerebral inactivity occurs during various brain disorders, including hypothermia, drug intoxication, long-lasting anoxia and brain trauma. It can also be induced in a therapeutic context, following the administration of high doses of barbiturate-derived compounds, to interrupt a hyper-refractory status epilepticus. Although altered sensory responses can be occasionally observed on an isoelectric electroencephalogram, the electrical membrane properties and synaptic responses of individual neurons during this cerebral state remain largely unknown. The aim of the present study was to characterize the intracellular correlates of a barbiturate-induced isoelectric electroencephalogram and to analyse the sensory-evoked synaptic responses that can emerge from a brain deprived of spontaneous electrical activity. We first examined the sensory responsiveness from patients suffering from intractable status epilepticus and treated by administration of thiopental. Multimodal sensory responses could be evoked on the flat electroencephalogram, including visually-evoked potentials that were significantly amplified and delayed, with a high trial-to-trial reproducibility compared to awake healthy subjects. Using an analogous pharmacological procedure to induce prolonged electro-cerebral inactivity in the rat, we could describe its cortical and subcortical intracellular counterparts. Neocortical, hippocampal and thalamo-cortical neurons were all silent during the isoelectric state and displayed a flat membrane potential significantly hyperpolarized compared with spontaneously active control states. Nonetheless, all recorded neurons could fire action potentials in response to intracellularly injected depolarizing current pulses and their specific intrinsic electrophysiological features were preserved. Manipulations of the membrane potential and intracellular injection of chloride in neocortical neurons failed to reveal an augmented synaptic inhibition during the isoelectric condition. Consistent with the sensory responses recorded from comatose patients, large and highly reproducible somatosensory-evoked potentials could be generated on the inactive electrocorticogram in rats. Intracellular recordings revealed that the underlying neocortical pyramidal cells responded to sensory stimuli by complex synaptic potentials able to trigger action potentials. As in patients, sensory responses in the isoelectric state were delayed compared to control responses and exhibited an elevated reliability during repeated stimuli. Our findings demonstrate that during prolonged isoelectric brain state neurons and synaptic networks are dormant rather than excessively inhibited, conserving their intrinsic properties and their ability to integrate and propagate environmental stimuli.

Building Up Absence Seizures in the Somatosensory Cortex: From Network to Cellular Epileptogenic Processes.

The epileptogenic processes leading to recurrent seizures in Genetic Epilepsies are largely unknown. Using the Genetic Absence Epilepsy Rat from Strasbourg, we investigated in vivo the network and single neuron mechanisms responsible for the early emergence of epileptic activity. Local field potential recordings in the primary somatosensory cortex (SoCx), from the second post-natal week to adulthood, showed that immature cortical discharges progressively evolved into typical spike-and-wave discharges following a 3-step maturation process. Intracellular recordings from deep-layer SoCx neurons revealed that this maturation was associated with an age-dependent increase in cortical neurons intrinsic excitability, combining a membrane depolarization and an enhancement of spontaneous firing rate with a leftward shift in their input-output relation. These cellular changes were accompanied by a progressive increase in the strength of the local synaptic activity associated with a growing propensity of neurons to generate synchronized oscillations. Chronic anti-absence treatment before the occurrence of mature cortical discharges did not alter epileptogenesis or the drug efficiency at adulthood. These findings demonstrate that recurrent absence seizures originate from the progressive acquisition of pro-ictogenic properties in SoCx neurons and networks during the post-natal period and that these processes cannot be interrupted by early anti-absence treatment.

Integrative properties and transfer function of cortical neurons initiating absence seizures in a rat genetic model.

KEY POINTS: Absence seizures are accompanied by spike-and-wave discharges in cortical electroencephalograms. These complex paroxysmal activities, affecting the thalamocortical networks, profoundly alter cognitive performances and preclude conscious perception. Here, using a well-recognized genetic model of absence epilepsy, we investigated in vivo how information processing was impaired in the ictogenic neurons, i.e. the population of cortical neurons responsible for seizure initiation. In between seizures, ictogenic neurons were more prone to generate bursting activity and their firing response to weak depolarizing events was considerably facilitated compared to control neurons. In the course of seizures, information processing became unstable in ictogenic cells, alternating between an increased and a decreased responsiveness to excitatory inputs, depending on the spike and wave patterns. The state-dependent modulation in the excitability of ictogenic neurons affects their inter-seizure transfer function and their time-to-time responsiveness to incoming inputs during absences. ABSTRACT: Epileptic seizures result from aberrant cellular and/or synaptic properties that can alter the capacity of neurons to integrate and relay information. During absence seizures, spike-and-wave discharges (SWDs) interfere with incoming sensory inputs and preclude conscious experience. The Genetic Absence Epilepsy Rats from Strasbourg (GAERS), a well-established animal model of absence epilepsy, allows exploration of the cellular basis of this impaired information processing. Here, by combining in vivo electrocorticographic and intracellular recordings from GAERS and control animals, we investigated how the pro-ictogenic properties of seizure-initiating cortical neurons modify their integrative properties and input-output operation during inter-ictal periods and during the spike (S-) and wave (W-) cortical patterns alternating during seizures. In addition to a sustained depolarization and an excessive firing rate in between seizures, ictogenic neurons exhibited a pronounced hyperpolarization-activated depolarization compared to homotypic control neurons. Firing frequency versus injected current relations indicated an increased sensitivity of GAERS cells to weak excitatory inputs, without modifications in the trial-to-trial variability of current-induced firing. During SWDs, the W-component resulted in paradoxical effects in ictogenic neurons, associating an increased membrane input resistance with a reduction in the current-evoked firing responses. Conversely, the collapse of cell membrane resistance during the S-component was accompanied by an elevated current-evoked firing relative to W-sequences, which remained, however, lower compared to inter-ictal periods. These findings show a dynamic modulation of ictogenic neurons' intrinsic properties that may alter inter-seizure cortical function and participate in compromising information processing in cortical networks during absences.

Excitability and responsiveness of rat barrel cortex neurons in the presence and absence of spontaneous synaptic activity in vivo.

The amplitude and temporal dynamics of spontaneous synaptic activity in the cerebral cortex vary as a function of brain states. To directly assess the impact of different ongoing synaptic activities on neocortical function, we performed in vivo intracellular recordings from barrel cortex neurons in rats under two pharmacological conditions generating either oscillatory or tonic synaptic drive. Cortical neurons membrane excitability and firing responses were compared, in the same neurons, before and after complete suppression of background synaptic drive following systemic injection of a high dose of anaesthetic. Compared to the oscillatory state, the tonic pattern resulted in a more depolarized and less fluctuating membrane potential (Vm), a lower input resistance (Rm) and steeper relations of firing frequency versus injected current (F-I). Whatever their temporal dynamics, suppression of background synaptic activities increased mean Vm, without affecting Rm, and induced a rightward shift of F-I curves. Both types of synaptic drive generated a high variability in current-induced firing rate and patterns in cortical neurons, which was much reduced after removal of spontaneous activity. These findings suggest that oscillatory and tonic synaptic patterns differentially facilitate the input-output function of cortical neurons but result in a similar moment-to-moment variability in spike responses to incoming depolarizing inputs.

High-fidelity multimode fibre-based endoscopy for deep brain in vivo imaging.

Progress in neuroscience relies on new techniques for investigating the complex dynamics of neuronal networks. An ongoing challenge is to achieve minimally invasive and high-resolution observations of neuronal activity in vivo inside deep brain areas. Recently introduced methods for holographic control of light propagation in complex media enable the use of a hair-thin multimode optical fibre as an ultranarrow imaging tool. Compared to endoscopes based on graded-index lenses or fibre bundles, this new approach offers a footprint reduction exceeding an order of magnitude, combined with a significant enhancement in resolution. We designed a compact and high-speed system for fluorescent imaging at the tip of a fibre, achieving a resolution of 1.18 ± 0.04 µm across a 50-µm field of view, yielding 7-kilopixel images at a rate of 3.5 frames/s. Furthermore, we demonstrate in vivo observations of cell bodies and processes of inhibitory neurons within deep layers of the visual cortex and hippocampus of anaesthetised mice. This study paves the way for modern microscopy to be applied deep inside tissues of living animal models while exerting a minimal impact on their structural and functional properties.

Induction of an Isoelectric Brain State to Investigate the Impact of Endogenous Synaptic Activity on Neuronal Excitability In Vivo.

The way neurons process information depends both on their intrinsic membrane properties and on the dynamics of the afferent synaptic network. In particular, endogenously-generated network activity, which strongly varies as a function of the state of vigilance, significantly modulates neuronal computation. To investigate how different spontaneous cerebral dynamics impact single neurons' integrative properties, we developed a new experimental strategy in the rat consisting in suppressing in vivo all cerebral activity by means of a systemic injection of a high dose of sodium pentobarbital. Cortical activities, continuously monitored by combined electrocorticogram (ECoG) and intracellular recordings are progressively slowed down, leading to a steady isoelectric profile. This extreme brain state, putting the rat into a deep comatose, was carefully monitored by measuring the physiological constants of the animal throughout the experiments. Intracellular recordings allowed us to characterize and compare the integrative properties of the same neuron embedded into physiologically relevant cortical dynamics, such as those encountered in the sleep-wake cycle, and when the brain was fully silent.