The spectral features of EEG responses to transcranial magnetic stimulation of the primary motor cortex depend on the amplitude of the motor evoked potentials.

Transcranial magnetic stimulation (TMS) of the primary motor cortex (M1) can excite both cortico-cortical and cortico-spinal axons resulting in TMS-evoked potentials (TEPs) and motor-evoked potentials (MEPs), respectively. Despite this remarkable difference with other cortical areas, the influence of motor output and its amplitude on TEPs is largely unknown. Here we studied TEPs resulting from M1 stimulation and assessed whether their waveform and spectral features depend on the MEP amplitude. To this aim, we performed two separate experiments. In experiment 1, single-pulse TMS was applied at the same supra-threshold intensity on primary motor, prefrontal, premotor and parietal cortices and the corresponding TEPs were compared by means of local mean field power and time-frequency spectral analysis. In experiment 2 we stimulated M1 at resting motor threshold in order to elicit MEPs characterized by a wide range of amplitudes. TEPs computed from high-MEP and low-MEP trials were then compared using the same methods applied in experiment 1. In line with previous studies, TMS of M1 produced larger TEPs compared to other cortical stimulations. Notably, we found that only TEPs produced by M1 stimulation were accompanied by a late event-related desynchronization (ERD-peaking at ~300 ms after TMS), whose magnitude was strongly dependent on the amplitude of MEPs. Overall, these results suggest that M1 produces peculiar responses to TMS possibly reflecting specific anatomo-functional properties, such as the re-entry of proprioceptive feedback associated with target muscle activation.

Stratification of unresponsive patients by an independently validated index of brain complexity.

OBJECTIVE: Validating objective, brain-based indices of consciousness in behaviorally unresponsive patients represents a challenge due to the impossibility of obtaining independent evidence through subjective reports. Here we address this problem by first validating a promising metric of consciousness-the Perturbational Complexity Index (PCI)-in a benchmark population who could confirm the presence or absence of consciousness through subjective reports, and then applying the same index to patients with disorders of consciousness (DOCs). METHODS: The benchmark population encompassed 150 healthy controls and communicative brain-injured subjects in various states of conscious wakefulness, disconnected consciousness, and unconsciousness. Receiver operating characteristic curve analysis was performed to define an optimal cutoff for discriminating between the conscious and unconscious conditions. This cutoff was then applied to a cohort of noncommunicative DOC patients (38 in a minimally conscious state [MCS] and 43 in a vegetative state [VS]). RESULTS: We found an empirical cutoff that discriminated with 100% sensitivity and specificity between the conscious and the unconscious conditions in the benchmark population. This cutoff resulted in a sensitivity of 94.7% in detecting MCS and allowed the identification of a number of unresponsive VS patients (9 of 43) with high values of PCI, overlapping with the distribution of the benchmark conscious condition. INTERPRETATION: Given its high sensitivity and specificity in the benchmark and MCS population, PCI offers a reliable, independently validated stratification of unresponsive patients that has important physiopathological and therapeutic implications. In particular, the high-PCI subgroup of VS patients may retain a capacity for consciousness that is not expressed in behavior. Ann Neurol 2016;80:718-729.

Shared reduction of oscillatory natural frequencies in bipolar disorder, major depressive disorder and schizophrenia.

INTRODUCTION: Recent studies have demonstrated that cortical brain areas tend to oscillate at a specific natural frequency when directly perturbed by transcranial magnetic stimulation (TMS). Fast electroencephalographic (EEG) oscillations, which typically originate from frontal regions, have been reported to be markedly reduced in schizophrenia. METHODS: Here we employed TMS/EEG to assess the natural frequency of the premotor area in a sample of 48 age-matched participants (12 each in major depression disorder (MDD)), bipolar disorder (BPD), schizophrenia (SCZ) and healthy controls. Event related spectral perturbations (ERSP) were obtained for each study participant using wavelet decomposition. RESULTS: TMS resulted in a significant activation of the beta/gamma band response (21-50 Hz) to frontal cortical perturbation in healthy control subjects. By contrast, the main frequencies of frontal EEG responses to TMS were significantly reduced in patients with BPD, MDD and SCZ (11-27 Hz) relative to healthy subjects. CONCLUSIONS: Patients with bipolar disorder, major depression and schizophrenia showed a significantly lower natural frequency of frontal cortico-thalamocortical circuits compared to healthy controls. These results suggest a common neurobiological mechanism of corticothalamic impairment. The most likely candidates include dysfunction of GABAergic circuits. LIMITATIONS: Further studies are needed to consider other biological markers, gene variants, and their interaction with clinical variables.

Bistability breaks-off deterministic responses to intracortical stimulation during non-REM sleep.

During non-rapid eye movement (NREM) sleep (stage N3), when consciousness fades, cortico-cortical interactions are impaired while neurons are still active and reactive. Why is this? We compared cortico-cortical evoked-potentials recorded during wakefulness and NREM by means of time-frequency analysis and phase-locking measures in 8 epileptic patients undergoing intra-cerebral stimulations/recordings for clinical evaluation. We observed that, while during wakefulness electrical stimulation triggers a chain of deterministic phase-locked activations in its cortical targets, during NREM the same input induces a slow wave associated with an OFF-period (suppression of power>20Hz), possibly reflecting a neuronal down-state. Crucially, after the OFF-period, cortical activity resumes to wakefulness-like levels, but the deterministic effects of the initial input are lost, as indicated by a sharp drop of phase-locked activity. These findings suggest that the intrinsic tendency of cortical neurons to fall into a down-state after a transient activation (i.e. bistability) prevents the emergence of stable patterns of causal interactions among cortical areas during NREM. Besides sleep, the same basic neurophysiological dynamics may play a role in pathological conditions in which thalamo-cortical information integration and consciousness are impaired in spite of preserved neuronal activity.

Assessing the effects of electroconvulsive therapy on cortical excitability by means of transcranial magnetic stimulation and electroencephalography.

Electroconvulsive therapy (ECT) has significant short-term antidepressant effects on drug-resistant patients with severe major depression. Animal studies have demonstrated that electroconvulsive seizures produce potentiation-like synaptic remodeling in both sub-cortical and frontal cortical circuits. However, the electrophysiological effects of ECT in the human brain are not known. In this work, we evaluated whether ECT induces a measurable change in the excitability of frontal cortical circuits in humans. Electroencephalographic (EEG) potentials evoked by transcranial magnetic stimulation (TMS) were collected before and after a course of ECT in eight patients with severe major depression. Cortical excitability was measured from the early and local EEG response to TMS. Clinical assessment confirmed the beneficial effects of ECT on depressive symptoms at the group level. TMS/EEG measurements revealed a clear-cut increase of frontal cortical excitability after ECT as compared to baseline, that was significant in each and every patient. The present findings corroborate in humans the idea that ECT may produce synaptic potentiation, as previously observed in animal studies. Moreover, results suggest that TMS/EEG may be employed in depressed patients to monitor longitudinally the electrophysiological effects of different therapeutic neuromodulators, e.g. ECT, repetitive TMS, and sleep deprivation. To the extent that depression involves an alteration of frontal cortical excitability, these measurements may be used to guide and evaluate treatment progression over time at the single-patient level.

Recovery of cortical effective connectivity and recovery of consciousness in vegetative patients.

Patients surviving severe brain injury may regain consciousness without recovering their ability to understand, move and communicate. Recently, electrophysiological and neuroimaging approaches, employing simple sensory stimulations or verbal commands, have proven useful in detecting higher order processing and, in some cases, in establishing some degree of communication in brain-injured subjects with severe impairment of motor function. To complement these approaches, it would be useful to develop methods to detect recovery of consciousness in ways that do not depend on the integrity of sensory pathways or on the subject's ability to comprehend or carry out instructions. As suggested by theoretical and experimental work, a key requirement for consciousness is that multiple, specialized cortical areas can engage in rapid causal interactions (effective connectivity). Here, we employ transcranial magnetic stimulation together with high-density electroencephalography to evaluate effective connectivity at the bedside of severely brain injured, non-communicating subjects. In patients in a vegetative state, who were open-eyed, behaviourally awake but unresponsive, transcranial magnetic stimulation triggered a simple, local response indicating a breakdown of effective connectivity, similar to the one previously observed in unconscious sleeping or anaesthetized subjects. In contrast, in minimally conscious patients, who showed fluctuating signs of non-reflexive behaviour, transcranial magnetic stimulation invariably triggered complex activations that sequentially involved distant cortical areas ipsi- and contralateral to the site of stimulation, similar to activations we recorded in locked-in, conscious patients. Longitudinal measurements performed in patients who gradually recovered consciousness revealed that this clear-cut change in effective connectivity could occur at an early stage, before reliable communication was established with the subject and before the spontaneous electroencephalogram showed significant modifications. Measurements of effective connectivity by means of transcranial magnetic stimulation combined with electroencephalography can be performed at the bedside while by-passing subcortical afferent and efferent pathways, and without requiring active participation of subjects or language comprehension; hence, they offer an effective way to detect and track recovery of consciousness in brain-injured patients who are unable to exchange information with the external environment.

Time-frequency spectral analysis of TMS-evoked EEG oscillations by means of Hilbert-Huang transform.

A single pulse of Transcranial Magnetic Stimulation (TMS) generates electroencephalogram (EEG) oscillations that are thought to reflect intrinsic properties of the stimulated cortical area and its fast interactions with other cortical areas. Thus, a tool to decompose TMS-evoked oscillations in the time-frequency domain on a millisecond timescale and on a broadband frequency range may help to understand information transfer across cortical oscillators. Some recent studies have employed algorithms based on the Wavelet Transform (WT) to study TMS-evoked EEG oscillations in healthy and pathological conditions. However, these methods do not allow to describe TMS-evoked EEG oscillations with high resolution in time and frequency domains simultaneously. Here, we first develop an algorithm based on Hilbert-Huang Transform (HHT) to compute statistically significant time-frequency spectra of TMS-evoked EEG oscillations on a single trial basis. Then, we compared the performances of the HHT-based algorithm with the WT-based one by applying both of them to a set of simulated signals. Finally, we applied both algorithms to real TMS-evoked potentials recorded in healthy or schizophrenic subjects. We found that the HHT-based algorithm outperforms the WT-based one in detecting the time onset of TMS-evoked oscillations in the classical EEG bands. These results suggest that the HHT-based algorithm may be used to study the communication between different cortical oscillators on a fine time scale.

EEG responses to TMS are sensitive to changes in the perturbation parameters and repeatable over time.

BACKGROUND: High-density electroencephalography (hd-EEG) combined with transcranial magnetic stimulation (TMS) provides a direct and non-invasive measure of cortical excitability and connectivity in humans and may be employed to track over time pathological alterations, plastic changes and therapy-induced modifications in cortical circuits. However, the diagnostic/monitoring applications of this technique would be limited to the extent that TMS-evoked potentials are either stereotypical (non-sensitive) or random (non-repeatable) responses. Here, we used controlled changes in the stimulation parameters (site, intensity, and angle of stimulation) and repeated longitudinal measurements (same day and one week apart) to evaluate the sensitivity and repeatability of TMS/hd-EEG potentials. METHODOLOGY/PRINCIPAL FINDINGS: In 10 volunteers, we performed 92 single-subject comparisons to evaluate the similarities/differences between pairs of TMS-evoked potentials recorded in the same/different stimulation conditions. For each pairwise comparison, we used non-parametric statistics to calculate a Divergence Index (DI), i.e., the percentage of samples that differed significantly, considering all scalp locations and the entire post-stimulus period. A receiver operating characteristic analysis showed that it was possible to find an optimal DI threshold of 1.67%, yielding 96.7% overall accuracy of TMS/hd-EEG in detecting whether a change in the perturbation parameters occurred or not. CONCLUSIONS/SIGNIFICANCE: These results demonstrate that the EEG responses to TMS essentially reflect deterministic properties of the stimulated neuronal circuits as opposed to stereotypical responses or uncontrolled variability. To the extent that TMS-evoked potentials are sensitive to changes and repeatable over time, they may be employed to detect longitudinal changes in the state of cortical circuits.

General indices to characterize the electrical response of the cerebral cortex to TMS.

Transcranial magnetic stimulation (TMS) combined with simultaneous high-density electroencephalography (hd-EEG) represents a straightforward way to gauge cortical excitability and connectivity in humans. However, the analysis, classification and interpretation of TMS-evoked potentials are hampered by scarce a priori knowledge about the physiological effect of TMS and by lack of an established data analysis framework. Here, we implemented a standardized, data-driven procedure to characterize the electrical response of the cerebral cortex to TMS by means of three synthetic indices: significant current density (SCD), phase-locking (PL) and significant current scattering (SCS). SCD sums up the amplitude of all significant currents induced by TMS, PL reflects the ability of TMS to reset the phase of ongoing cortical oscillations, while SCS measures the average distance of significantly activated sources from the site of stimulation. These indices are aimed at capturing different aspects of brain responsiveness, ranging from global cortical excitability towards global cortical connectivity. We analyzed the EEG responses to TMS of Brodmann's area 19 at increasing intensities in five healthy subjects. The spatial distribution and time course of SCD, PL and SCS revealed a reproducible profile of excitability and connectivity, characterized by a local activation threshold around a TMS-induced electric field of 50 V/m and by a selective propagation of TMS-evoked activation from occipital to ipsilateral frontal areas that reached a maximum at 70-100 ms. These general indices may be used to characterize the effects of TMS on any cortical area and to quantitatively evaluate cortical excitability and connectivity in physiological and pathological conditions.

Natural frequencies of human corticothalamic circuits.

The frequency tuning of a system can be directly determined by perturbing it and by observing the rate of the ensuing oscillations, the so called natural frequency. This approach is used, for example, in physics, in geology, and also when one tunes a musical instrument. In the present study, we employ transcranial magnetic stimulation (TMS) to directly perturb a set of selected corticothalamic modules (Brodmann areas 19, 7, and 6) and high-density electroencephalogram to measure their natural frequency. TMS consistently evoked dominant alpha-band oscillations (8-12 Hz) in the occipital cortex, beta-band oscillations (13-20 Hz) in the parietal cortex, and fast beta/gamma-band oscillations (21-50 Hz) in the frontal cortex. Each cortical area tended to preserve its own natural frequency also when indirectly engaged by TMS through brain connections and when stimulated at different intensities, indicating that the observed oscillations reflect local physiological mechanisms. These findings were reproducible across individuals and represent the first direct characterization of the coarse electrophysiological properties of three associative areas of the human cerebral cortex. Most importantly, they indicate that, in healthy subjects, each corticothalamic module is normally tuned to oscillate at a characteristic rate. The natural frequency can be directly measured in virtually any area of the cerebral cortex and may represent a straightforward and flexible way to probe the state of human thalamocortical circuits at the patient's bedside.

Augmentative repetitive navigated transcranial magnetic stimulation (rTMS) in drug-resistant bipolar depression.

OBJECTIVES: The efficacy of transcranial magnetic stimulation (TMS) has been poorly investigated in bipolar depression. The present study aimed to assess the efficacy of low-frequency repetitive TMS (rTMS) of the right dorsolateral prefrontal cortex (DLPFC) combined with brain navigation in a sample of bipolar depressed subjects. METHODS: Eleven subjects with bipolar I or bipolar II disorder and major depressive episode who did not respond to previous pharmacological treatment were treated with three weeks of open-label rTMS at 1 Hz, 110% of motor threshold, 300 stimuli/day. RESULTS: All subjects completed the trial showing a statistically significant improvement on the 21-item Hamilton Depression Rating Scale (HAM-D), Montgomery-Asberg Depression Rating Scale, and Clinical Global Impression severity of illness scale (ANOVAs with repeated measures: F = 22.36, p < 0.0001; F = 12.66, p < 0.0001; and F = 10.41, p < 0.0001, respectively). In addition, stimulation response, defined as an endpoint HAM-D score reduction of > or =50% compared to baseline, was achieved by 6 out of 11 subjects, 4 of whom were considered remitters (HAM-D endpoint score < or = 8). Partial response (endpoint HAM-D score reduction between 25% and 50%) was achieved by 3/11 patients. No manic/hypomanic activation was detected during the treatment according to Young Mania Rating Scale scores (ANOVAs with repeated measures: F = 0.62, p = 0.61). Side effects were slight and were limited to the first days of treatment. CONCLUSIONS: Augmentative low-frequency rTMS of the right DLPFC combined with brain navigation was effective and well tolerated in a small sample of drug-resistant bipolar depressive patients, even though the lack of a sham controlled group limits confidence in the results.

Behavioral and degeneration changes in the basal forebrain systems of aged rats: a quantitative study in the region of the basal forebrain after levo-acetyl-carnitine treatments assessed by Abercrombie estimation.

One group of six male control rats [21 months old] and one group of six male rats of the same age, singularly stored in a cage, and treated with acetyl-l-carnitine-HCl (ALCAR: 60 mg/kg/day/p.o.) for six months were tested in the spatial learning/memory Morris maze-water task and for atrophy and cell loss in seven myelo- and cytostructurally defined basal forebrain (BF) cholinergic regions [Gritti et al., 1993 J Comp Neurol 329: 438-457]. Coronal sections 25 mum thick were cut through the BF regions and processed every 200 mum for choline acetyltransferase (ChAT) immunohistochemistry. The ALCAR-treated rats had significantly shorter exit times on the Morris maze-water task test than the control rats (ANOVA-enzyme: F(1,39)=112.5, P=0.0001; sessions: F(3,39)=10.41, P=0.0001; interaction: F(3,39)=5.09, P=0.0044). Degenerative morphological changes in the BF ChAT-positive cells were observed in the control rats, but not in the treated animals, in: the diagonal band of Broca, the magnocellular preoptic nucleus, the olfactory tubercle, the substantia innominata, and the globus pallidus (ANOVA-enzyme: F(1,2)=14, P=0,0003; structures: F(6,7)=4, P=0,0018; interaction: F(6,7)=3, P=0,0043). In the diagonal band of Broca (P<0.0494) and in the magnocellular preoptic nucleus (P<0.0117) there were significantly fewer ChAT-positive neurons in the aged control rats than in the ALCAR-treated rats. These results demonstrate that in rats aged from 15 to 21 months ALCAR treatment significantly attenuated spatial learning/memory impairment on the Morris maze-water task and also importantly reduced the degeneration in size and number of cholinergic cells in the BF.

Sleep, but not febrile responses of Fisher 344 rats to immune challenge are affected by aging.

Sleep is altered in response to infection and immune challenge in humans and non-human animals. Although there are changes in sleep and facets of immune function with aging, sleep responses of aged subjects to immune challenge have received little, if any attention. To test the hypothesis that aging affects sleep responses to immune challenge, intracerebroventricular injections of interleukin 1 (IL-1) were given to young and aged rats and subsequent sleep-wake behavior was determined. Under basal conditions and in the absence of an immune challenge, sleep patterns of young (3 months) and aged (25-27 months) Fisher 344 rats did not differ. In young animals, IL-1 (2.5 ng) enhanced non-rapid eye movement (NREM) sleep, inhibited rapid eye movement (REM) sleep, and induced fever. In aged animals, IL-1 administration did not alter NREM sleep, but REM sleep was inhibited and brain temperature increased to the same extent observed in young animals. These results show that alterations in sleep following immune challenge are impacted by aging, whereas febrile responses are not. Since it has been postulated that enhanced NREM sleep may facilitate recovery from microbial infection, the present results also suggest that the lack of NREM sleep responses of aged rats to immune challenge may contribute to the increased infection-induced morbidity and mortality of aged organisms.

Interleukin-1beta enhances non-rapid eye movement sleep when microinjected into the dorsal raphe nucleus and inhibits serotonergic neurons in vitro.

Interleukin-1 (IL-1) and IL-1 receptors are constitutively expressed in normal brain. IL-1 increases non-rapid eye movements (NREM) sleep in several animal species, an effect mediated in part by interactions with the serotonergic system. The site(s) in brain at which interactions between IL-1 and the serotonergic system increase NREM sleep remain to be identified. The dorsal raphe (DRN) is the origin of the major ascending serotonergic pathways to the forebrain, and it contains IL-1 receptors. This study examined the hypothesis that IL-1 increases NREM sleep by acting at the level of the DRN. IL-1beta (0.25 and 0.5 ng) was microinjected into the DRN of freely behaving rats and subsequent effects on sleep-wake activity were determined. IL-1beta 0.5 ng increased NREM sleep during the first 2 h post-injection from 33.5 +/- 3.7% after vehicle microinjection to 42.9 +/- 3.0% of recording time. To determine the effects of IL-1beta on electrophysiological properties of DRN serotonergic neurons, intracellular recordings were performed in a guinea-pig brain stem slice preparation. In 26 of 32 physiologically and pharmacologically identified serotonergic neurons, IL-1beta superfusion (25 ng/mL) decreased spontaneous firing rates by 50%, from 1.6 +/- 0.2 Hz (before IL-1beta superfusion) to 0.8 +/- 0.2 Hz. This effect was reversible upon washout. These results show that IL-1beta increases NREM sleep when administered directly into the DRN. Serotonin enhances wakefulness and these novel data also suggest that IL-1beta-induced enhancement of NREM sleep could be due in part to the inhibition of DRN serotonergic neurons.

EEG slow (approximately 1 Hz) waves are associated with nonstationarity of thalamo-cortical sensory processing in the sleeping human.

Intracellular studies reveal that, during slow wave sleep (SWS), the entire cortical network can swing rhythmically between extremely different microstates, ranging from wakefulness-like network activation to functional disconnection in the space of a few hundred milliseconds. This alternation of states also involves the thalamic neurons and is reflected in the EEG by a slow (<1 Hz) oscillation. These rhythmic changes, occurring in the thalamo-cortical circuits during SWS, may have relevant, phasic effects on the transmission and processing of sensory information. However, brain reactivity to sensory stimuli, during SWS, has traditionally been studied by means of sequential averaging, a procedure that necessarily masks any short-term fluctuation of responsiveness. The aim of this study was to provide a dynamic evaluation of brain reactivity to sensory stimuli in naturally sleeping humans. To this aim, single-trial somatosensory evoked potentials (SEPs) were grouped and averaged as a function of the phase of the ongoing sleep slow (<1 Hz) oscillation. This procedure revealed a dynamic profile of responsiveness, which was conditioned by the phase of the spontaneous sleep EEG. Overall, the amplitude of the evoked potential changed sistematically, increasing and approaching wakefulness levels along the negative slope of the EEG oscillation and decaying below SWS average levels along the positive drift. These marked and fast changes of stimulus-correlated electrical activity involved both short (N20) and long latency (P60 and P100) components of SEPs. In addition, the observed short-term response variability appeared to be centrally generated and specifically related to the evolution of the spontaneous oscillatory pattern. The present findings demonstrate that thalamo-cortical processing of sensory information is not stationary in the very short period (approximately 500 ms) during natural SWS.

Awakening from sleep.

Awakening is a crucial event for the organism. The transition from sleep to waking implies physiological processes which lead to a new behavioural state. Spontaneous awakenings have varying features which may change as a function of several factors. The latter include intrasleep architecture, circadian phase, time awake, age, or disordered sleep. Despite its clear theoretical and clinical importance, the topic of awakening (in humans) has received little attention so far. This contribution focuses on major issues which relate to awakening from both basic (experimental) and clinical research. Recent knowledge on neurophysiological mechanisms is reported. The experimental data which provide in the human suggestions on the regulation of awakening are discussed, mainly those concerning sleep architecture and homeostatic/circadian factors also in a life-span perspective, since age is a powerful factor which may influence awakening. Clinical contributions will examine two main sleep disorders: insomnia and hypersomnia. Daytime functioning is shown in insomniac patients and compared to other pathologies like sleep apnea. A final section evokes links between some types of night waking and psychological factors.