**6. Neurophysiological techniques**

The neurophysiological approach to patients with DOC allows the recording of electrical activities of both CNS and Peripheral Nervous System (PNS) and provides a functional assessment, which can be integrated with data obtained mainly from morphological neuroimaging techniques (CT and MRI). The combined use of various neurophysiological examinations, such as Electroencephalogram (EEG), Evoked Related Potentials (ERPs), Transcranial Magnetic Stimulation (TMS), Deep Brain Stimulation (DBS), EEG in association with fMRI, contributes to the topographic and functional diagnosis of the various anatomical structures of the injured CNS and the PNS. The electrophysiological signals recorded from electrodes placed on the surface of the scalp reflect spatially the average postsynaptic potential originated by large neuronal populations. Experimental evidence and clinical observations suggest functional correlations among the neural mechanisms of sensory information, cognitive performance, sleep-wake cycle, alertness and electrophysiological signals generated by neuronal activity. A greater amount of patients can be assessed with electrophysiological techniques, including those who may not have access to a MRI due to geographic, financial, or physical (i.e., metal plates or pins) impairment.

The electroencephalogram in patients in VS has shown a spectrum of abnormalities with changes during the wake-sleep cycle. Patterns have included delta and theta activity and

97% specific for predicting an unfavorable outcome after 1 year, compared with 85% for DTI and 75% for 1H-MRS. Similarly, sensitivity was better with the combined analysis (86%)

To study metabolite changes from a wider area of the brain, with the advantage of identifying more anatomical and functional details, a few investigators have used 1H-MRSI (Holshouser et al., 2006; Marino et al. 2007; Shutter et al., 2006; Signoretti et al., 2002, 2008). This technique has an advantage over single-voxel 1H-MRS because generates individual spectra from multiple voxels at the same time. Also 1H-MRSI studies have highlighted close

Neurometabolite concentrations obtained soon after injury may be useful for predicting individual outcome. The decrease of NAA and the increase of Lac, seen by Marino et al. (2007) early after brain injury, were correlated with GOS score. Then these 1H-MRS data

However it is need to note that 1H-MRS studies in patients with DOC are heterogeneous in terms of patient nature, injury types, time from cerebral damage, voxel location, methods and timing outcome assessment. In addition, in many studies the metabolite concentrations were expressed in terms of semiquantitative ratios. The assumption that the concentration of Cr as reference metabolite remains constant may be incorrect, especially in acute conditions. It is therefore advisable to obtain concentration expressed in standard units by applying absolute quantification. Some studies have expressed metabolite concentrations in term of absolute quantification (Brook et al., 2000; Friedman et al., 1999; Marino et al. 2007; Ross et

Data reported so far demonstrate that MRS measure have the potential to provide new and important biological brain markers able to predict clinical outcome, helping in the therapeutic interventions, clinical and rehabilitative management of these patients, as well

The neurophysiological approach to patients with DOC allows the recording of electrical activities of both CNS and Peripheral Nervous System (PNS) and provides a functional assessment, which can be integrated with data obtained mainly from morphological neuroimaging techniques (CT and MRI). The combined use of various neurophysiological examinations, such as Electroencephalogram (EEG), Evoked Related Potentials (ERPs), Transcranial Magnetic Stimulation (TMS), Deep Brain Stimulation (DBS), EEG in association with fMRI, contributes to the topographic and functional diagnosis of the various anatomical structures of the injured CNS and the PNS. The electrophysiological signals recorded from electrodes placed on the surface of the scalp reflect spatially the average postsynaptic potential originated by large neuronal populations. Experimental evidence and clinical observations suggest functional correlations among the neural mechanisms of sensory information, cognitive performance, sleep-wake cycle, alertness and electrophysiological signals generated by neuronal activity. A greater amount of patients can be assessed with electrophysiological techniques, including those who may not have access to a MRI due to geographic, financial, or physical (i.e., metal plates or pins)

The electroencephalogram in patients in VS has shown a spectrum of abnormalities with changes during the wake-sleep cycle. Patterns have included delta and theta activity and

than with either DTI (79%) or MRS (75%).

al., 2000; Shutter et al., 2004).

impairment.

as to assist with family education.

**6. Neurophysiological techniques** 

correlation between metabolite alterations and potential recovery.

may be, at this stage, a reliable index of injury severity and disease outcome.

spindle and alpha-like rhythms, but they are more diffusely distributed than in the typical posterior regions and are not reactive to sound, pain, and light stimuli (Chokroverty, 1975; Huges, 1978). During sleep, fewer muscle twitches are observed, but a REM sleep remains (Oksenberg et al., 2001). In most patients, the transition from wakefulness to sleep is accompanied by some desynchronization of the background activity. Very-low-voltage EEG activity is all that can be detected in some patients. In others, persistent alpha activity is the most remarkable feature. In around 10% of patients with VS, the EEG is nearly normal late in the course of disease but without evidence of vision-induced alpha blocking (Danze et al., 1989). There have been occasional reports of isoelectric EEGs in patients in a VS, although it has not been confirmed (Higashi et al., 1977; Mizrahi et al., 1985). Typical epileptiform activity is unusual in patients in VS, as seizure activity is (The Multi-Society Task Force on PVS, 1994). Clinical recovery from the vegetative state may be paralleled by diminished delta and theta activity and reappearance of reactive alpha rhythm. Indeed, Babiloni (2009) has observed that occipital source power in the alpha band (8-13 Hz) of resting EEG, when calculated with low-resolution electromagnetic tomography (LORETA), is correlated with recovery outcome at 3-month follow-up in a group of VS patients; those who made a behavioural recovery had higher resting alpha band power than those who did not make a significant recovery.

The EEG in MCS shows diffuse slowing brain activity, mainly of the theta band, and in most cases responsive to external stimuli. However, there are insufficient data as well as the typical pattern of MCS concerns. Evoked potentials have been studied in patients in a VS and showed normal brainstem auditory responses but abnormal somatosensory responses: prolonged conduction time or absence of scalp potentials. ERPs are more useful than EEG in the differential diagnosis between VS and MCS. ERPs studies focusing on the assessment of conscious awareness have frequently examined four specific components: the N100, the mismatch negativity (MMN), the P300, and the N400 (Connoly & D'Arcy, 2000).

In a recent work, the authors focused on the prediction of consciousness recovery in patients with post-traumatic VS. They used a classical two-stimulus oddball task to elicit the P300 using the patient's own name as deviant and a pure tone as standard stimulus (''subject's own name" paradigm). There is evidence that the amplitude of the P300 wave increases when more salient stimuli are used, such as the own first name instead of visual or auditory deviants. The authors found that P300 is a strong predictor of future recovery of consciousness in VS. This finding is in line with several studies that have confirmed the utility of P300 evoked by deviant tones to predict awakening and favourable outcome from coma and VS (Cavinato et al., 2009). In another study Cavinato et al., (2011) continue to using the "subject's own name" paradigm, but add a pure tone and an "other first name" paradigm. The authors instructed their patients to count the occurrence of deviant stimuli to better differentiate between patients in VS and MCS. The study indicates that in 6 out of 11 patients fulfilling the behavioral criteria for VS a reliable P300 component could be observed in all two conditions. These findings corroborate earlier reports showing that 38% of patients in VS generate a P300 wave. The patients in MCS exhibit significantly longer P300 latencies for the "subject's own name" and the "other first name" paradigms than patients in VS. The increase of P300 latencies for more complex and salient paradigms in MCS but not in VS might help in the difficult differential diagnosis of MCS vs. VS.

The TMS, for high temporal resolution, was proposed as an additional functional imaging technique for the study of cognitive function. To date only some studies have assessed VS and MCS patients with TMS. Moosavi et al. (1999) applied TMS to the hand and leg motor

Neuroimaging and Outcome Assessment in Vegetative and Minimally Conscious State 193

In this paragraph we try to identify some markers of consciousness with prognostic value, based on literature review. As it has been clearly established in clinical practice, significant spontaneous recovery frequently occurs during the subacute period (Wilson et al., 2002; Giacino & Trott 2004). Two factors can facilitate cognitive recovery of these patients: young

Literature findings demonstrated that no clinical tools strongly predicted good outcome. In contrast, complementary examinations such as electrophysiological and functional neuroimaging studies objectively measure residual brain functions and are indicative of

Several studies explore the prognostic validity of behavioural assessment scales (i.e. GOS, Coma Recovery Scale, Wessex Head Injury Matrix, Western Neuro Sensory Stimulation Profile, DRS, Functional Independence Measure), electrophysiological measures (ERPs), and functional neuroimaging (PET, fMRI), to predict outcome in patients with low levels of consciousness. Particular progress towards addressing this objective has been made using brain imaging techniques such as PET and fMRI. Schiff et al. (2002) suggested that rather than a complete loss of cortical function some patients retain "island" of preserved cognitive functions. PET and fMRI studies suggest that a higher-level associative cortical activation seems to predict recovery of consciousness with a 93% specificity and 69% sensitivity (Di et al., 2008). PET work has identified preserved responses to a variety of sensory stimuli, including photographs of familiar people, noxious, tactile (Laureys et al. 2000; Owen et al., 2002; Boly et al., 2004) in some vegetative and minimally conscious patients. Some studies underline the importance of the cognitive ERPs in the assessment of residual functions in comatose, VS, or MCS patients. As a general rule, early ERPs (such as the absence of cortical response on somatosensory evoked-potentials) predict bad outcome, while cognitive ERPs are indicative of recovery of consciousness (Vanhaudenhuyse et al., 2007). Moreover, auditory cognitive ERPs are useful to investigate residual cognitive functions, such as echoic memory (Mismatch Negativity), acoustical and semantic discrimination (P300), and incongruent language detection (N400). In VS patients, cognitive potential are more frequently obtained when using stimuli that are more ecologic or have an emotional content

age and immediate medical assistance after the injury.

(such as the patients' own name) than when using classical sine tones.

Electrophysiological and functional neuroimaging studies may provide useful and objective information to the outcome and possibly cognition of patients with low levels of consciousness (Di et al., 2007). To date, there have been no detailed studies of these patients combining and correlating specific neuropsychological tools and functional imaging, in order to evaluate the cognitive changes and recovery over time (Bekinschtein et al, 2005). In spite of the important findings, functional neuroimaging cannot, and should not replace, clinical and behavioural evaluation as the criterion standard for assessment of patients with DOC. It offers an objective method of differentiating brain activity measured at rest and during external stimulation, but further studies are needed to assess the temporal evolution of individual patients' somatosensory and cognitive processing (Giacino et al., 2006). Despite converging agreement about the definition of persistent vegetative state, recent reports have raised concerns about the accuracy of diagnosis in some patients, and the extent to which, in a selection of cases, residual cognitive functions may remain undetected. Objective assessment of residual cognitive function can be extremely difficult as motor responses may be minimal, inconsistent, and difficult to document in many patients, or may be undetectable in others because no cognitive output is possible (Owen et al., 2002). There are no standards of care to guide the selection of rehabilitation assessment and treatment

recovery of consciousness.

area in 19 patients, few months after severe anoxic brain injury. Eleven patients were in VS, while eight patients were in MCS. The VS patient group differed from the MCS patient group in having a higher threshold, longer duration, and greater irregularity in the form of the response, while the threshold, form, and latency of motor evoked potentials (MEPs) from the MCS group were similar to healthy control subjects. In another study, TMS is used to monitor recovery. The authors examined MEPs from upper and lower limbs in 27 patients in the subacute period and then at 6 and 12 months post – ictus. During the study period, the authors observed an overall trend toward an increase and decrease of latency of MEPs. MEPs from upper and lower limbs progressively normalized in all patients, and at one year after trauma, only 12% of patients had mild abnormalities in MEP responses (Mazzini et al., 1999).

TMS elicited MEP responses in the majority of severely brain damage patients, and a trend toward an increase of amplitude and decrease of latency of MEPs could be observed during the recovery period.

DBS works on reactivating the cortex, aiming to produce a functional recovery. In study of Yamamoto et al. (2010) patients in VS were treated with DBS. Eight of the patients recovered from VS and were able to obey verbal commands at 13 and 10 months in the case of head trauma and a year and a half in the case of vascular disease after comatose brain injury, and no patients without DBS recovered from VS spontaneously within 24 months after brain injury.

In the last years the interest in using of neurophysiological investigations (EEG and EPs) in association with fMRI, has grown: the combination of these different neuroimaging techniques allows study of different components of the brain's activity (e.g., neurovascular coupling, electromagnetic activity) with both a high temporal and spatial resolution (Gosseries et al., 2008).

Clinical neurophysiology procedures are useful as easily performed, non invasive and repeatable at the bed side. These methods provide irreplaceable data about the degree of neuronal dysfunctions and their evolution, and gave also information to assess the outcome.
