367
Restorative Neurology and Neuroscience 32 (2014) 367–379 DOI 10.3233/RNN-130323 IOS Press
Bihemispheric tDCS enhances language recovery but does not alter BDNF levels in chronic aphasic patients Paola Marangoloa,b,∗ , Valentina Fiorib , Francesca Gelfob,c , Jacob Shofanyb , Carmelina Razzanob , Carlo Caltagironeb,c and Francesco Angeluccib,∗ a Facolt` a
di Medicina, Universit`a Politecnica Marche, Ancona, Italy Fondazione Santa Lucia, Roma, Italy c Dipartimento di Medicina dei Sistemi, Universit` a degli studi di Roma “Tor Vergata”, Roma, Italy b IRCCS
Abstract. Purpose: Several studies have shown that transcranial direct current stimulation (tDCS) is a useful tool to enhance language recovery in aphasia. It has also been suggested that modulation of the neurotrophin brain-derived neurotrophic factor (BDNF) might be part of the mechanisms involved in tDCS effects on synaptic connectivity. However, all language studies have previously investigated the effects using unihemispheric stimulation. The purpose of the present study is to investigate the role of bihemispheric tDCS on language recovery and BDNF serum levels. Methods: Seven aphasic persons underwent an intensive language therapy in two different conditions: real bihemispheric stimulation over the left and right Broca’s areas and a sham condition. Results: After the stimulation, patients exibited a significant recovery in three language tasks (picture description, noun and verb naming) compared to the sham condition which persisted in the follow-up session. No significant differences were found in BDNF serum levels after tDCS stimulation and in the follow-up session. However, a significant positive correlation was present for the real stimulation condition between percent changes in BDNF levels and in the verb naming task. Conclusions: The data suggest that this novel approach may potentiate the recovery of language in chronic aphasia. They also emphasize the importance to further investigate the role of possible biomarkers associated with tDCS treatment response in language recovery. Keywords: Aphasia, bihemispheric tDCS, Broca’s area, language recovery, BDNF
1. Introduction Transcranial direct current stimulation (tDCS) is a non-invasive brain stimulation technique that has garnered increasing interest due to its modulatory effect on cognitive functions and motor behaviour in healthy ∗ Corresponding author: Prof. Paola Marangolo, Department of Experimental and Clinical Medicine, Universit`a Politecnica delle Marche, Ancona – Italia. Tel.: +39 071 2206093; Fax: +39 071 2206214; E-mail: [email protected]; Dott. Francesco Angelucci (should be referred for BDNF data), IRCCS Santa Lucia Foundation, Department of Clinical and Behavioural Neurology, 00179 Rome, Italy. Tel.: +39 06 51501550; Fax: +39 06 51501552; E-mail: [email protected].
and brain-damaged subjects (Fl¨oel et al., 2008; Bastani and Jaberzadeh, 2012; Grefkes and Fink, 2012; Holland and Crinion, 2012; Sandrini et al., 2012; Feng et al., 2013; Schulz et al., 2013). During tDCS, weak polarizing direct currents are delivered to the cortex via two electrodes placed on the scalp. The nature of the effect depends on the polarity of the current. Generally, the anode increases cortical excitability when applied over the region of interest with the cathode above the contralateral orbit or above the shoulder (as the reference electrode), whereas the cathode decreases it, limiting the resting membrane potential (Nitsche and Paulus, 2011).
0922-6028/14/$27.50 © 2014 – IOS Press and the authors. All rights reserved
368
P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
A number of studies have already emphasized tDCS role as an adjuvant strategy to improve word-finding difficulties in language aphasic disorders. Indeed, it has been demonstrated that left stroke patients exhibit greater recovery of word-retrieval deficits when the language treatment was coupled with repeated tDCS stimulation (Baker et al., 2010; Fiori et al., 2011; Fridriksson et al., 2011; Kang et al., 2011; Marangolo et al., 2013a; see Monti et al., 2013 for a review). In previous works, we found beneficial effects in word production after one week of intensive language treatment together with 20 minutes of tDCS stimulation (Fiori et al., 2011; Marangolo et al., 2013a). More recently, evidence for the efficacy of bihemispheric tDCS stimulation has been also produced (Vines et al., 2008; Lindenberg et al., 2010; Lefebvre et al., 2012; Mordillo-Mateos et al., 2012; Fusco et al., 2013). This was based on the assumption that the stimulation over the perilesional areas together with the suppression of the contralateral cortex might reduce the inhibition exerted by the unaffected hemisphere via the transcallosal pathway leading to a better recovery (Ziemann, 2005; Kim et al., 2006; Schlaug et al., 2008). Supporting this idea, bihemispheric tDCS and simultaneous physical/occupational therapy given over 5 consecutive sessions significantly improved motor function in a group of twenty chronic stroke patients when compared to the sham group (Lindenberg et al., 2010). However, despite its increasing use in experimental and clinical settings, the cellular and molecular mechanisms underlying tDCS remain unknown. Some authors have hypothesized that anodic (A) stimulation elicits a prolonged increment in cortical plasticity, probably due to the depolarization of the neuronal membrane and changes in the synaptic connections of the N-methyl-d-aspartate (NMDA) receptors involved in long-term potentiation (LTP, Stagg and Nitsche, 2011; Monte-Silva et al., in press). This physiological effect, LTP and its opposite, long-term depression (LTD), has been mostly characterized in ex vivo animal hippocampal slices (Ranieri et al., 2012). The effect of repeated applications of A-tDCS in restoring function after stroke appears to be associated with an increased LTP-like plasticity in the neocortex. This effect is dependent on modulation of cortical interneurons rather than pyramidal neuron excitability, and pharmacological studies indicate that function at noradrenergic, dopaminergic, serotoninergic, GABAergic and glutamatergic synapses may all influence the dura-
tion and sometimes the magnitude of the effects of A-tDCS (Nitsche et al., 2004). It has also been suggested that the effect of A-tDCS on synaptic plasticity may also depend on other molecular pathways, such as the secretion of brain-derived neurotrophic factor (BDNF), a protein belonging to the neurotrophin family and widely expressed in various parts of the human brain (Murer et al., 1999; Zolads and Pilc, 2010). BDNF is able to regulate synaptic plasticity by inducing functional and structural changes (McAllister et al., 1999; Gomez-Pinilla et al., 2008; Gottmann et al., 2009). At present, the impact of brain stimulation techniques on BDNF has been mainly investigated using repetitive transcranial stimulation (rTMS). In animal models, rTMS has been shown to produce opposite effects on hippocampal and cortical BDNF levels of awake or anesthetized rats, respectively increasing or decreasing them (Gersner et al., 2011). In humans, Angelucci et al. (2004) compared the effect of low and high frequency rTMS delivered on the motor cortex in a group of ten healthy subjects and four amyotrophic lateral sclerosis (ALS) patients on BDNF plasma levels. BDNF levels were progressively decreased by 1 Hz rTMS in healthy subjects, while, there was no effect of 1 Hz rTMS on BDNF in ALS patients. High frequency rTMS determined a transitory decrease in BDNF plasma levels. Cumulatively these findings suggest that rTMS might infuence the BDNF production by interfering with neuronal activity. However, Gedge et al. (2012) compared pre- and post-treatment serum BDNF levels in patients with drug- resistant major depressive disorder (MDD) who received rTMS and found that this treatment did not significantly alter BDNF concentrations (see also Lang et al., 2006; 2008). To date, only three studies have explored the impact of BDNF on tDCS effects. Antal et al. (2010) showed in a retrospective analysis that carriers of the Val66Met BDNF polymorphism enhanced tDCS-induced plasticity as compared to Val66Val carriers. In contrast, Cheeran et al. (2008) found no differences between these groups for cathodic tDCS-induced plasticity. Di Lazzaro and colleagues (2012) explored the impact of a long-lasting suppression of motor cortical excitability induced by prolonged cathodic tDCS (20 min of stimulation) on BDNF levels. Cortical excitability to single and pairedpulse TMS was measured both for the stimulated and contralateral hemiphere, before and up to 24 h after 20 min of cathodic tDCS. Although tDCS exerted a pronounced suppression of MEP amplitude that was still
P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
significant at 3 h after the end of stimulation, the BDNF genotype was not associated with tDCS after-effects. Taken together, these findings suggest that the effect of brain stimulation on BDNF levels may vary according to the type of stimulation, the brain area stimulated and the protocol used. As far as we know, until now no reports have studied the effect of bihemispheric tDCS on language recovery and BDNF serum levels. In a very recent study, our group (Marangolo et al., 2013b) have shown that bihemispheric tDCS (delivered over the frontal areas) coupled with intensive language therapy (10 consecutive sessions) significantly improved articulatory difficulties in a group of eight chronic aphasics as compared to the sham group. Thus, in the present study we investigated whether the same group of aphasic patients (except one) would further benefit by bihemispheric tDCS stimulation in association to a different language protocol aimed at the recovery of informative speech. Moreover, we investigated the possibility that the effect of bihemispheric tDCS on language recovery is associated to changes in BDNF serum level.
2. Materials and methods 2.1. Participants Seven participants (5 men and 2 female) who had suffered a single left hemisphere ischemic stroke were included in the study (see Fig. 1). Inclusion criteria for the study were native Italian proficiency, premorbid right handedness (based on the Edinburgh Handedness Questionnaire, Oldfield, 1971), a single left hemispheric stroke at least 6 months prior to the investigation, and no acute or chronic neurological symptoms requiring medication. The data analyzed in the current study were collected in accordance with the Helsinky Declaration and the Institutional Review Board of the IRCCS Fondazione Santa Lucia, Rome, Italy. Prior to participation, all patients signed informed consent forms. 2.2. Clinical data The seven patients were diagnosed with severe non-fluent aphasia because of their reduced spontaneous speech with frequent word-finding difficulties. All patients were administered a standardized language test including different language tasks (Esame
369
del Linguaggio II, cut-off 100%, Ciurli et al., 1996). In a picture description task, their language production was telegraphic, with verb omissions and not well-formed sentences. Verbs and nouns naming were impaired. Their comprehension abilities were functionally adequate at the language test, while they still had difficulties in more complex auditory comprehension tasks (Token test, cut-off 29/36, De Renzi and Faglioni, 1978, see Table 1). 3. Procedure 3.1. Transcranial direct current stimulation (tDCS) tDCS was applied using a battery driven Eldith (neuroConn GmbH) Programmable Direct Current Stimulator with a pair of surface-soaked sponge electrodes (5 cm × 7 cm). If applied according to safety guidelines, tDCS is considered to be a safe brain stimulation technique with minor adverse effects (Poreisz et al., 2007). Real stimulation consisted of 20 minutes of 2 mA direct current with the anode placed over the ipsilesional and the cathode over the contralesional inferior frontal gyrus (IFG) (F5 and F7 of the extended International 10–20 system for EEG electrode placement). For sham stimulation, the same electrode positions were used. The current was ramped up to 2 mA and slowly decreased over 30 seconds to ensure the typical initial tingling sensation (Gandiga et al., 2006). In both conditions (real vs. sham), patients underwent concurrent speech therapy (see below). For each condition, the language treatment was performed in ten daily sessions (Monday-Friday, weekend off, Monday-Friday). There was 14-day intersession interval between the real and the sham conditions. The order of conditions was randomized across subjects (see Fig. 2). Both the patient and the clinician were blinded with respect to the administration of tDCS which was applied by a third person who had nothing to do with the study. At the end of each condition, subjects were asked if they were aware of which condition (real or sham) they were in. None of the subjects was able to ascertain differences in intensity of sensation between the two conditions. 3.2. Language treatment At the beginning (baseline, T0), at the end (T10), and at one week after the end of each treatment
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P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
371
Table 1 Sociodemographic and Clinical data of the seven non fluent aphasic subjects. For each language task, the percentage of correct responses are reported (Esame del Linguaggio, cut-off score 100%, Ciurli et al., 1996) Participant
Sex
Age
Ed. level
1 2 3 4 5 6 7
M M M F F F M
60 53 49 60 62 51 68
8 13 18 8 13 13 5
Time post-onset
Picture descrip.
Noun naming
Verb naming
Word comprehension
Sentence comprehension
Token test
0 0 0 0 0 0 0
0 0 47 20 5 15 0
0 0 0 0 0 0 0
100 100 100 100 100 100 100
60 90 80 100 75 90 100
11/36 13/36 12/36 10/36 9/36 14/36 10/36
2 years, 6 months 10 months 3 years 1 year, 2 months 6 years, 2 months 6 months 4 years, 8 months
condition (follow-up, F/U), all patients were administered the language tests. During the administration of the language test, the experimenter was blinded to the condition of the subject. All patients underwent an intensive, one hour and a half, daily language treatment using a pragmatic approach (Basso, 2010; Marangolo, 2010). During the treatment both the patient and the therapist set up a natural verbal exchange of salient information on different cartoons story (i.e.,
the “Flower pot”, Huber and Gleber, 1982; and the “Quarrel”, Nicholas and Brookshire, 1993). The therapist had to accept all the information provided by the patient and tried to relate it to the topic of conversation in order to improve its content and informativeness. The goal of the therapy was to make the patient more informative day-by-day with the context and to bring him/her to talk about the story without the therapist’s support.
Fig. 1. Lesion descriptions for each aphasic patient (L: Left, R: Right). The figure shows the MRI acquisitions of all patients. The left panels display the sagittal view of the damaged left hemisphere; the central panels show the coronal view of the brain; the right panels show the spared right hemisphere. Patients 1, 2, 4 underwent a 1.5 T scanner exam (Siemens Vision Magnetom MR system) with acquisition of a T1 3D sequence for patient 1 and 2 and a clinical T1 sequence for patient 4. Patients 3, 5, 6, 7 underwent a 3.0 T scanner exam (Phillips Achieva Magnetom MR system) with acquisition of a T1 3D sequence. Pt 1. lesion is localized in the left fronto-temporo-parieto-occipital cortex. At frontal level the damage involves the middle frontal gyrus and the inferior frontal gyrus (Broca’s area). At temporo-parietal level the damage partially spared the temporal pole and involves the full extension of the superior temporal gyrus (Wernicke’s area), the middle temporal gyrus, the angular and the supramarginal gyri and the inferior parietal lobule. At occipital level the damage involves the superior and the middle occipital gyri. The lesion also includes the insula. At sub-cortical level the damage partially involves the homolateral sub-cortical nuclei (putamen and globus pallidus), the external and internal capsule. Pt 2. lesion is localized in the left fronto-temporo-parietal cortices. At frontal level the damage laterally involves the inferior frontal gyrus (Broca’s area), the middle frontal gyrus, the superior frontal gyrus and the pre-central gyrus, and medially the medial frontal gyrus and the anterior cingulate gyrus. At temporal level the lesion includes the pole as well as the superior (Wernicke’s area) and the middle temporal gyrus. At parietal level the damage involves the post-central gyrus and the inferior parietal lobule. The lesion also includes the insula. At sub-cortical level the lesion includes the putamen and the globus pallidus, the external and internal capsule. Pt. 3. is localized in the left fronto-temporo-parietal cortex. At frontal level the damage involves the middle frontal gyrus, the inferior frontal gyrus (Broca’s area), part of the lateral frontal pole and of the pre-central gyrus. At temporal level the damage includes the temporal pole, the full extension of the superior temporal gyrus (Wernicke’s area), and most part of the middle temporal gyrus. At parietal level the damage involves most part of the post-central gyrus, of the inferior parietal lobule and of the angular and supramarginal gyri. The lesion also includes the insula. At sub-cortical level the lesion includes the putamen and the globus pallidus, the external and internal capsule. Pt 4. lesion is localized in the left fronto-parietal cortex. At frontal level the damage involves the inferior frontal gyrus (Broca’s area), the middle frontal gyrus and the pre-central gyrus. At parietal level the damage includes the post-central gyrus and part of the inferior parietal lobule. The lesion also involves the insula. At sub-cortical level the lesion includes the thalamus, the putamen and the globus pallidus, the caudate nucleus, the external and internal capsule. Pt 5. is localized in the left fronto-temporo-parietal cortex. At frontal level the damage involves the middle frontal gyrus and the inferior frontal gyrus (Broca’s area). At temporal-parietal level the damage includes the temporal pole, the full extension of the superior temporal gyrus (Wernicke’s area), the middle and most part of the inferior temporal gyrus and of the angular and supramarginal gyri. The lesion also includes the insula. At sub-cortical level the lesion involves the thalamus, the putamen and the globus pallidus, the caudate nucleus and the external and internal capsule. Pt 6. is localized in the left fronto-temporo-parietal cortex. At frontal level the damage involves the middle frontal gyrus, the inferior frontal gyrus (Broca’s area), part of the lateral frontal pole and the pre-central gyrus. At temporal level the damage includes part of the temporal pole and the full extension of the superior temporal gyrus (Wernicke’s area). At the parietal level the damage involves most of the post-central gyrus, the inferior parietal lobule, the angular and part of the supramarginal gyri. The lesion also includes the insula. At sub-cortical level the lesion involves the putamen and the globus pallidus, the caudate nucleus, the external and the internal capsule. Pt 7. A cortical-subcortical atrophy involving the fronto-parietal cortex was present. The lesion is localized in the left insula. At sub-cortical level the lesion involves the thalamus, the putamen and the globus pallidus, the caudate nucleus, the external and internal capsule.
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Fig. 2. Overview of study design: at the beginning (T0), at the end (T10), and at follow-up (F/U), all patients were administered the language tests and their blood samples were collected. All patients underwent bihemispheric stimulation (20 min, 2 mA) over the frontal areas during language treatment. Each condition (real vs. sham) was performed in ten consecutive daily sessions (Monday-Friday, weekend off, Monday-Friday) with 14 days of intersession interval. The order of conditions was randomized across subjects.
3.3. Blood sampling In all participants, in both experimental conditions (real vs. sham), blood samples were collected before the treatment (T0), at the end of treatment (T10) and at one week after the end of treatment (F/U). Venous blood was collected into sampling tubes that were centrifuged within 20 minutes after sampling at 2000× g for 20 minutes. Serum was then aliquoted and stored at −80◦ C until analysis. 3.4. Determination of BDNF content in serum BDNF was detected in sandwich ELISAs according to the manufacturer’s instructions (R and D Systems, Minneapolis, MN, USA). This sandwich ELISA is set in order to measure natural and recombinant human mature BDNF in cell culture medium and serum. All assays were performed on F-bottom 96-well plates (Nunc, Wiesbaden, Germany). Tertiary antibody was conjugated to horseradish peroxidase. Wells were developed with tetramethylbenzidine and measured at 450/570 nm. BDNF content was quantified against a standard curve calibrated with known amounts of protein. The detection limits were
Restorative Neurology and Neuroscience 32 (2014) 367–379 DOI 10.3233/RNN-130323 IOS Press
Bihemispheric tDCS enhances language recovery but does not alter BDNF levels in chronic aphasic patients Paola Marangoloa,b,∗ , Valentina Fiorib , Francesca Gelfob,c , Jacob Shofanyb , Carmelina Razzanob , Carlo Caltagironeb,c and Francesco Angeluccib,∗ a Facolt` a
di Medicina, Universit`a Politecnica Marche, Ancona, Italy Fondazione Santa Lucia, Roma, Italy c Dipartimento di Medicina dei Sistemi, Universit` a degli studi di Roma “Tor Vergata”, Roma, Italy b IRCCS
Abstract. Purpose: Several studies have shown that transcranial direct current stimulation (tDCS) is a useful tool to enhance language recovery in aphasia. It has also been suggested that modulation of the neurotrophin brain-derived neurotrophic factor (BDNF) might be part of the mechanisms involved in tDCS effects on synaptic connectivity. However, all language studies have previously investigated the effects using unihemispheric stimulation. The purpose of the present study is to investigate the role of bihemispheric tDCS on language recovery and BDNF serum levels. Methods: Seven aphasic persons underwent an intensive language therapy in two different conditions: real bihemispheric stimulation over the left and right Broca’s areas and a sham condition. Results: After the stimulation, patients exibited a significant recovery in three language tasks (picture description, noun and verb naming) compared to the sham condition which persisted in the follow-up session. No significant differences were found in BDNF serum levels after tDCS stimulation and in the follow-up session. However, a significant positive correlation was present for the real stimulation condition between percent changes in BDNF levels and in the verb naming task. Conclusions: The data suggest that this novel approach may potentiate the recovery of language in chronic aphasia. They also emphasize the importance to further investigate the role of possible biomarkers associated with tDCS treatment response in language recovery. Keywords: Aphasia, bihemispheric tDCS, Broca’s area, language recovery, BDNF
1. Introduction Transcranial direct current stimulation (tDCS) is a non-invasive brain stimulation technique that has garnered increasing interest due to its modulatory effect on cognitive functions and motor behaviour in healthy ∗ Corresponding author: Prof. Paola Marangolo, Department of Experimental and Clinical Medicine, Universit`a Politecnica delle Marche, Ancona – Italia. Tel.: +39 071 2206093; Fax: +39 071 2206214; E-mail: [email protected]; Dott. Francesco Angelucci (should be referred for BDNF data), IRCCS Santa Lucia Foundation, Department of Clinical and Behavioural Neurology, 00179 Rome, Italy. Tel.: +39 06 51501550; Fax: +39 06 51501552; E-mail: [email protected].
and brain-damaged subjects (Fl¨oel et al., 2008; Bastani and Jaberzadeh, 2012; Grefkes and Fink, 2012; Holland and Crinion, 2012; Sandrini et al., 2012; Feng et al., 2013; Schulz et al., 2013). During tDCS, weak polarizing direct currents are delivered to the cortex via two electrodes placed on the scalp. The nature of the effect depends on the polarity of the current. Generally, the anode increases cortical excitability when applied over the region of interest with the cathode above the contralateral orbit or above the shoulder (as the reference electrode), whereas the cathode decreases it, limiting the resting membrane potential (Nitsche and Paulus, 2011).
0922-6028/14/$27.50 © 2014 – IOS Press and the authors. All rights reserved
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P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
A number of studies have already emphasized tDCS role as an adjuvant strategy to improve word-finding difficulties in language aphasic disorders. Indeed, it has been demonstrated that left stroke patients exhibit greater recovery of word-retrieval deficits when the language treatment was coupled with repeated tDCS stimulation (Baker et al., 2010; Fiori et al., 2011; Fridriksson et al., 2011; Kang et al., 2011; Marangolo et al., 2013a; see Monti et al., 2013 for a review). In previous works, we found beneficial effects in word production after one week of intensive language treatment together with 20 minutes of tDCS stimulation (Fiori et al., 2011; Marangolo et al., 2013a). More recently, evidence for the efficacy of bihemispheric tDCS stimulation has been also produced (Vines et al., 2008; Lindenberg et al., 2010; Lefebvre et al., 2012; Mordillo-Mateos et al., 2012; Fusco et al., 2013). This was based on the assumption that the stimulation over the perilesional areas together with the suppression of the contralateral cortex might reduce the inhibition exerted by the unaffected hemisphere via the transcallosal pathway leading to a better recovery (Ziemann, 2005; Kim et al., 2006; Schlaug et al., 2008). Supporting this idea, bihemispheric tDCS and simultaneous physical/occupational therapy given over 5 consecutive sessions significantly improved motor function in a group of twenty chronic stroke patients when compared to the sham group (Lindenberg et al., 2010). However, despite its increasing use in experimental and clinical settings, the cellular and molecular mechanisms underlying tDCS remain unknown. Some authors have hypothesized that anodic (A) stimulation elicits a prolonged increment in cortical plasticity, probably due to the depolarization of the neuronal membrane and changes in the synaptic connections of the N-methyl-d-aspartate (NMDA) receptors involved in long-term potentiation (LTP, Stagg and Nitsche, 2011; Monte-Silva et al., in press). This physiological effect, LTP and its opposite, long-term depression (LTD), has been mostly characterized in ex vivo animal hippocampal slices (Ranieri et al., 2012). The effect of repeated applications of A-tDCS in restoring function after stroke appears to be associated with an increased LTP-like plasticity in the neocortex. This effect is dependent on modulation of cortical interneurons rather than pyramidal neuron excitability, and pharmacological studies indicate that function at noradrenergic, dopaminergic, serotoninergic, GABAergic and glutamatergic synapses may all influence the dura-
tion and sometimes the magnitude of the effects of A-tDCS (Nitsche et al., 2004). It has also been suggested that the effect of A-tDCS on synaptic plasticity may also depend on other molecular pathways, such as the secretion of brain-derived neurotrophic factor (BDNF), a protein belonging to the neurotrophin family and widely expressed in various parts of the human brain (Murer et al., 1999; Zolads and Pilc, 2010). BDNF is able to regulate synaptic plasticity by inducing functional and structural changes (McAllister et al., 1999; Gomez-Pinilla et al., 2008; Gottmann et al., 2009). At present, the impact of brain stimulation techniques on BDNF has been mainly investigated using repetitive transcranial stimulation (rTMS). In animal models, rTMS has been shown to produce opposite effects on hippocampal and cortical BDNF levels of awake or anesthetized rats, respectively increasing or decreasing them (Gersner et al., 2011). In humans, Angelucci et al. (2004) compared the effect of low and high frequency rTMS delivered on the motor cortex in a group of ten healthy subjects and four amyotrophic lateral sclerosis (ALS) patients on BDNF plasma levels. BDNF levels were progressively decreased by 1 Hz rTMS in healthy subjects, while, there was no effect of 1 Hz rTMS on BDNF in ALS patients. High frequency rTMS determined a transitory decrease in BDNF plasma levels. Cumulatively these findings suggest that rTMS might infuence the BDNF production by interfering with neuronal activity. However, Gedge et al. (2012) compared pre- and post-treatment serum BDNF levels in patients with drug- resistant major depressive disorder (MDD) who received rTMS and found that this treatment did not significantly alter BDNF concentrations (see also Lang et al., 2006; 2008). To date, only three studies have explored the impact of BDNF on tDCS effects. Antal et al. (2010) showed in a retrospective analysis that carriers of the Val66Met BDNF polymorphism enhanced tDCS-induced plasticity as compared to Val66Val carriers. In contrast, Cheeran et al. (2008) found no differences between these groups for cathodic tDCS-induced plasticity. Di Lazzaro and colleagues (2012) explored the impact of a long-lasting suppression of motor cortical excitability induced by prolonged cathodic tDCS (20 min of stimulation) on BDNF levels. Cortical excitability to single and pairedpulse TMS was measured both for the stimulated and contralateral hemiphere, before and up to 24 h after 20 min of cathodic tDCS. Although tDCS exerted a pronounced suppression of MEP amplitude that was still
P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
significant at 3 h after the end of stimulation, the BDNF genotype was not associated with tDCS after-effects. Taken together, these findings suggest that the effect of brain stimulation on BDNF levels may vary according to the type of stimulation, the brain area stimulated and the protocol used. As far as we know, until now no reports have studied the effect of bihemispheric tDCS on language recovery and BDNF serum levels. In a very recent study, our group (Marangolo et al., 2013b) have shown that bihemispheric tDCS (delivered over the frontal areas) coupled with intensive language therapy (10 consecutive sessions) significantly improved articulatory difficulties in a group of eight chronic aphasics as compared to the sham group. Thus, in the present study we investigated whether the same group of aphasic patients (except one) would further benefit by bihemispheric tDCS stimulation in association to a different language protocol aimed at the recovery of informative speech. Moreover, we investigated the possibility that the effect of bihemispheric tDCS on language recovery is associated to changes in BDNF serum level.
2. Materials and methods 2.1. Participants Seven participants (5 men and 2 female) who had suffered a single left hemisphere ischemic stroke were included in the study (see Fig. 1). Inclusion criteria for the study were native Italian proficiency, premorbid right handedness (based on the Edinburgh Handedness Questionnaire, Oldfield, 1971), a single left hemispheric stroke at least 6 months prior to the investigation, and no acute or chronic neurological symptoms requiring medication. The data analyzed in the current study were collected in accordance with the Helsinky Declaration and the Institutional Review Board of the IRCCS Fondazione Santa Lucia, Rome, Italy. Prior to participation, all patients signed informed consent forms. 2.2. Clinical data The seven patients were diagnosed with severe non-fluent aphasia because of their reduced spontaneous speech with frequent word-finding difficulties. All patients were administered a standardized language test including different language tasks (Esame
369
del Linguaggio II, cut-off 100%, Ciurli et al., 1996). In a picture description task, their language production was telegraphic, with verb omissions and not well-formed sentences. Verbs and nouns naming were impaired. Their comprehension abilities were functionally adequate at the language test, while they still had difficulties in more complex auditory comprehension tasks (Token test, cut-off 29/36, De Renzi and Faglioni, 1978, see Table 1). 3. Procedure 3.1. Transcranial direct current stimulation (tDCS) tDCS was applied using a battery driven Eldith (neuroConn GmbH) Programmable Direct Current Stimulator with a pair of surface-soaked sponge electrodes (5 cm × 7 cm). If applied according to safety guidelines, tDCS is considered to be a safe brain stimulation technique with minor adverse effects (Poreisz et al., 2007). Real stimulation consisted of 20 minutes of 2 mA direct current with the anode placed over the ipsilesional and the cathode over the contralesional inferior frontal gyrus (IFG) (F5 and F7 of the extended International 10–20 system for EEG electrode placement). For sham stimulation, the same electrode positions were used. The current was ramped up to 2 mA and slowly decreased over 30 seconds to ensure the typical initial tingling sensation (Gandiga et al., 2006). In both conditions (real vs. sham), patients underwent concurrent speech therapy (see below). For each condition, the language treatment was performed in ten daily sessions (Monday-Friday, weekend off, Monday-Friday). There was 14-day intersession interval between the real and the sham conditions. The order of conditions was randomized across subjects (see Fig. 2). Both the patient and the clinician were blinded with respect to the administration of tDCS which was applied by a third person who had nothing to do with the study. At the end of each condition, subjects were asked if they were aware of which condition (real or sham) they were in. None of the subjects was able to ascertain differences in intensity of sensation between the two conditions. 3.2. Language treatment At the beginning (baseline, T0), at the end (T10), and at one week after the end of each treatment
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P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
371
Table 1 Sociodemographic and Clinical data of the seven non fluent aphasic subjects. For each language task, the percentage of correct responses are reported (Esame del Linguaggio, cut-off score 100%, Ciurli et al., 1996) Participant
Sex
Age
Ed. level
1 2 3 4 5 6 7
M M M F F F M
60 53 49 60 62 51 68
8 13 18 8 13 13 5
Time post-onset
Picture descrip.
Noun naming
Verb naming
Word comprehension
Sentence comprehension
Token test
0 0 0 0 0 0 0
0 0 47 20 5 15 0
0 0 0 0 0 0 0
100 100 100 100 100 100 100
60 90 80 100 75 90 100
11/36 13/36 12/36 10/36 9/36 14/36 10/36
2 years, 6 months 10 months 3 years 1 year, 2 months 6 years, 2 months 6 months 4 years, 8 months
condition (follow-up, F/U), all patients were administered the language tests. During the administration of the language test, the experimenter was blinded to the condition of the subject. All patients underwent an intensive, one hour and a half, daily language treatment using a pragmatic approach (Basso, 2010; Marangolo, 2010). During the treatment both the patient and the therapist set up a natural verbal exchange of salient information on different cartoons story (i.e.,
the “Flower pot”, Huber and Gleber, 1982; and the “Quarrel”, Nicholas and Brookshire, 1993). The therapist had to accept all the information provided by the patient and tried to relate it to the topic of conversation in order to improve its content and informativeness. The goal of the therapy was to make the patient more informative day-by-day with the context and to bring him/her to talk about the story without the therapist’s support.
Fig. 1. Lesion descriptions for each aphasic patient (L: Left, R: Right). The figure shows the MRI acquisitions of all patients. The left panels display the sagittal view of the damaged left hemisphere; the central panels show the coronal view of the brain; the right panels show the spared right hemisphere. Patients 1, 2, 4 underwent a 1.5 T scanner exam (Siemens Vision Magnetom MR system) with acquisition of a T1 3D sequence for patient 1 and 2 and a clinical T1 sequence for patient 4. Patients 3, 5, 6, 7 underwent a 3.0 T scanner exam (Phillips Achieva Magnetom MR system) with acquisition of a T1 3D sequence. Pt 1. lesion is localized in the left fronto-temporo-parieto-occipital cortex. At frontal level the damage involves the middle frontal gyrus and the inferior frontal gyrus (Broca’s area). At temporo-parietal level the damage partially spared the temporal pole and involves the full extension of the superior temporal gyrus (Wernicke’s area), the middle temporal gyrus, the angular and the supramarginal gyri and the inferior parietal lobule. At occipital level the damage involves the superior and the middle occipital gyri. The lesion also includes the insula. At sub-cortical level the damage partially involves the homolateral sub-cortical nuclei (putamen and globus pallidus), the external and internal capsule. Pt 2. lesion is localized in the left fronto-temporo-parietal cortices. At frontal level the damage laterally involves the inferior frontal gyrus (Broca’s area), the middle frontal gyrus, the superior frontal gyrus and the pre-central gyrus, and medially the medial frontal gyrus and the anterior cingulate gyrus. At temporal level the lesion includes the pole as well as the superior (Wernicke’s area) and the middle temporal gyrus. At parietal level the damage involves the post-central gyrus and the inferior parietal lobule. The lesion also includes the insula. At sub-cortical level the lesion includes the putamen and the globus pallidus, the external and internal capsule. Pt. 3. is localized in the left fronto-temporo-parietal cortex. At frontal level the damage involves the middle frontal gyrus, the inferior frontal gyrus (Broca’s area), part of the lateral frontal pole and of the pre-central gyrus. At temporal level the damage includes the temporal pole, the full extension of the superior temporal gyrus (Wernicke’s area), and most part of the middle temporal gyrus. At parietal level the damage involves most part of the post-central gyrus, of the inferior parietal lobule and of the angular and supramarginal gyri. The lesion also includes the insula. At sub-cortical level the lesion includes the putamen and the globus pallidus, the external and internal capsule. Pt 4. lesion is localized in the left fronto-parietal cortex. At frontal level the damage involves the inferior frontal gyrus (Broca’s area), the middle frontal gyrus and the pre-central gyrus. At parietal level the damage includes the post-central gyrus and part of the inferior parietal lobule. The lesion also involves the insula. At sub-cortical level the lesion includes the thalamus, the putamen and the globus pallidus, the caudate nucleus, the external and internal capsule. Pt 5. is localized in the left fronto-temporo-parietal cortex. At frontal level the damage involves the middle frontal gyrus and the inferior frontal gyrus (Broca’s area). At temporal-parietal level the damage includes the temporal pole, the full extension of the superior temporal gyrus (Wernicke’s area), the middle and most part of the inferior temporal gyrus and of the angular and supramarginal gyri. The lesion also includes the insula. At sub-cortical level the lesion involves the thalamus, the putamen and the globus pallidus, the caudate nucleus and the external and internal capsule. Pt 6. is localized in the left fronto-temporo-parietal cortex. At frontal level the damage involves the middle frontal gyrus, the inferior frontal gyrus (Broca’s area), part of the lateral frontal pole and the pre-central gyrus. At temporal level the damage includes part of the temporal pole and the full extension of the superior temporal gyrus (Wernicke’s area). At the parietal level the damage involves most of the post-central gyrus, the inferior parietal lobule, the angular and part of the supramarginal gyri. The lesion also includes the insula. At sub-cortical level the lesion involves the putamen and the globus pallidus, the caudate nucleus, the external and the internal capsule. Pt 7. A cortical-subcortical atrophy involving the fronto-parietal cortex was present. The lesion is localized in the left insula. At sub-cortical level the lesion involves the thalamus, the putamen and the globus pallidus, the caudate nucleus, the external and internal capsule.
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P. Marangolo et al. / Bihemispheric tDCS enhances language recovery
Fig. 2. Overview of study design: at the beginning (T0), at the end (T10), and at follow-up (F/U), all patients were administered the language tests and their blood samples were collected. All patients underwent bihemispheric stimulation (20 min, 2 mA) over the frontal areas during language treatment. Each condition (real vs. sham) was performed in ten consecutive daily sessions (Monday-Friday, weekend off, Monday-Friday) with 14 days of intersession interval. The order of conditions was randomized across subjects.
3.3. Blood sampling In all participants, in both experimental conditions (real vs. sham), blood samples were collected before the treatment (T0), at the end of treatment (T10) and at one week after the end of treatment (F/U). Venous blood was collected into sampling tubes that were centrifuged within 20 minutes after sampling at 2000× g for 20 minutes. Serum was then aliquoted and stored at −80◦ C until analysis. 3.4. Determination of BDNF content in serum BDNF was detected in sandwich ELISAs according to the manufacturer’s instructions (R and D Systems, Minneapolis, MN, USA). This sandwich ELISA is set in order to measure natural and recombinant human mature BDNF in cell culture medium and serum. All assays were performed on F-bottom 96-well plates (Nunc, Wiesbaden, Germany). Tertiary antibody was conjugated to horseradish peroxidase. Wells were developed with tetramethylbenzidine and measured at 450/570 nm. BDNF content was quantified against a standard curve calibrated with known amounts of protein. The detection limits were
