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Cb1 receptor antagonism/inverse agonism increases motor system excitability in humans

NEUPSY-10386; No of Pages 9 CB1 receptor antagonism/inverse agonism increasesmotor system excitability in humans A. Oliviero A. Arevalo-Martin , M. Rotondi , D. García-Ovejero L. Mordillo-Mateos , A. Lozano-Sicilia , I. Panyavin , L. Chiovato ,J. Aguilar G. Foffani V. Di Lazzaro , E. Molina-Holgado a FENNSI Group, Hospital Nacional de Parapléjicos, SESCAM, Toledo, Spainb Laboratory of Neuroinflammation, Unidad de Neurologia Experimental, Hospital Nacional de Parapléjicos, SESCAM,Toledo, Spainc Unit of Internal Medicine and Endocrinology, Fondazione Salvatore Maugeri I.R.C.C.S., University of Pavia, Pavia, Italyd Neurosignals Group, Hospital Nacional de Parapléjicos, SESCAM, Toledo, Spaine Institute of Neurology, Università Cattolica, Rome, Italy Received 9 December 2010; received in revised form 14 March 2011; accepted 17 April 2011 CB1 receptor is highly expressed in cerebral structures related to motor control, such as motor cortex, basal ganglia and cerebellum. In the spinal cord, the expression of CB1 receptors has also been observed in ventral motor neurons, interneurons and primary afferents, i.e., in the cells that may be part of the circuits involved in motor control. It is known that the antagonist/inverse agonist of CB1 receptors Rimonabant penetrates the blood–brain barrier and produces a broadrange of central psychoactive effects in humans. Based on the occurrence of central effects inhumans treated with Rimonabant and on the location of CB1 receptors, we hypothesized that theapplication of Rimonabant can also affect the motor system. We tested the effects of a singledose of 20 mg of Rimonabant on the excitability of motor cortex and of spinal motor neurons inorder to detect a possible drug action on motor system at cortical and spinal levels. For thispurpose we use classical protocols of transcranial magnetic and electrical stimulation (TMS andTES). Single and paired pulse TMS and TES were used to assess a number of parameters of corticalinhibition and cortical excitability as well as of the excitability of spinal motor neurons. Wedemonstrated that a single oral dose of 20 mg of Rimonabant can increase motor systemexcitability at cortical and spinal levels. This opens new avenues to test the CB1R antagonists/inverse agonists for the treatment of a number of neurological dysfunctions in which can be ⁎ Corresponding author at: FENNSI Group, Hospital Nacional de Parapléjicos, SESCAM, Finca La Peraleda s/n, 45071, Toledo, Spain. Tel.: +34 925396831; fax: + 34 925247745.
E-mail address: (A. Oliviero).
0924-977X/$ - see front matter 2011 Elsevier B.V. All rights reserved.
doi: Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), doi: A. Oliviero et al.
useful to increase the excitability levels of motor system. Virtually all the disorderscharacterized by a reduced output of the motor cortex can be included in the list of thedisorders that can be treated using CB1 antagonists/reverse agonists (e.g. stroke, traumaticbrain injury, spinal cord injury, multiple sclerosis, fatigue syndromes, parkinsonisms, etc.).
2011 Elsevier B.V. All rights reserved.
ceptor can be found at inhibitory or excitatory synapses. In thespinal cord, CB1 is distributed at high levels in the dorsal horn, Cannabinoids are lipophylic molecules that were first de- lamina X, dorsolateral funiculus, and in lower levels in motor scribed as the psychoactive constituents of marijuana. The neurons of the ventral horn and astrocytes (see discovery of the specific receptors, the endogenous ligands for a review). In lampreys, an animal model extensively for these receptors and the enzymatic machinery for their used to study locomotor networks, CB1 modulates the balance synthesis and degradation, revealed the existence of a whole between excitation and inhibition to modify the frequency of endocannabinoid system (To date, two locomotor rhythm ( cannabinoid receptors have been identified and cloned: CB1, Based on the occurrence of central effects in humans which is mainly expressed in the central nervous system treated with Rimonabant and on the location of CB1 (CNS), and CB2, preferentially located in the immune receptors, we hypothesized that the application of Rimona- system cells. Arachidonoyl ethanolamide (anandamide) and bant can also affect the motor system. The aim of the 2-arachidonoylglycerol (2-AG) are the two major endocan- present study is to test the effects of a single dose of 20 mg of nabinoids. These molecules are not stored in vesicles but Rimonabant on the excitability of motor cortex and of spinal produced from membrane lipid precursors in response to motor neurons in order to detect a possible drug action on calcium influx. This characteristic together with the fact motor system at cortical and spinal levels. For this purpose that CB1 receptor is mainly expressed on presynaptic axons we use classical protocols of transcranial magnetic and and the synthesizing machinery is expressed on the post- electrical stimulation (TMS and TES). Our results show that sinaptic axons make the endocannabinoid synthesis an on CB1 blockade produces a significant increase in motor circuit demand retrograde signaling for the regulation of CNS neuronal excitability and synaptic plasticity ).
In the brain, CB1 receptor is highly expressed in cerebral structures related to motor control, such as motor cortex, basal ganglia and cerebellum ). In the spinal cord, the expression of CB1 Nine healthy volunteers (all males, mean age 32.5 ± 4.8 years) receptors has also been observed in ventral motor neurons, participated in TMS experiments. None of the subjects had been interneurons and primary afferents, i.e., in the cells that may treated with neuroactive drugs in the 60 days prior to participating be part of the circuits involved in motor control in this electrophysiological study, which was performed in accor- dance with the Declaration of Helsinki and approved by the local ethics committee. A subgroup of six subjects participated in the on these data, several groups have studied the effects of experiments with transcranial electrical stimulation.
targeting the endocannabinoid system in animals to treat Rimonabant, a selective CB1 receptor blocker, was the first motor dysfunction, observing amelioration of hyperkinetic medication of its kind to become available commercially, in 2006.
dysfunctions and spasticity with CB1 agonists and enhance- However, due to adverse effects, such as nausea, diarrhea, dizziness ment of motor activity in hypokinetic disorders with CB1 and psychiatric side effects (i.e., anxiety, depression, agitation,increased suicide rate, etc.) ( antagonists ().
) Rimonabant was officially withdrawn from the market by the Moreover, cannabinoids have been shown to modulate spinal European Medicines Agency (EMEA) in January 2009. Our experi- ments were performed during the end of 2007/beginning of 2008, before any recommendations about the drug suspension and long before EMEA approval of Rimonabant was officially withdrawn.
It is known that Rimonabant (5-(4-Chlorophenyl)-1-(2,4- dichloro-phenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide) penetrates the blood–brain barrier and, at 2.2. Experimental setup doses used to obtain its metabolic effects (20 mg per day),produces a broad range of central psychoactive effects in Motor cortex excitability parameters were measured at baseline (before drug intake) and at three time points after drug intake. Thethree time points evaluated after the Rimonabant administration ). Being an antagonist/inverse agonist of CB1 cannabinoid were 2.5 h (plasma peak concentration, according to the datasheet receptors, the mechanism of action of Rimonabant on neuronal of Sanofi Aventis' Acomplia), 24 h and 48 h. Drug was taken orally activity depends on the cellular expression of CB1 receptor.
as a single oral dose (Rimonabant 20 mg). Rimonabant effects on CB1 is preferentially distributed in presynaptic elements, vigilance were moderate and did not interfere with subjects' ability where CB1 activation modulates voltage gated Ca2+ channels to fully comply with the requirements of the experimental protocol.
or K+ channels inhibiting neurotransmitter release. CB1 re- Hitherto, three subjects experienced agitation and anxiety (this Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), Effects of a CB1 antagonist on cortical excitability effect lasted an average of 6 h) and one experienced nausea preceded the test stimulus of 2, 3, 4, 5, 10, 15, and 25 ms). Each interval was pseudorandomly repeated and intermixed with the teststimuli (six conditions with only the test stimulus repeated 5 times).
2.3. Single pulse transcranial magnetic and electric The amplitude of the conditioned responses at the four differentinhibitory ISIs and the amplitude of the conditioned responses at the stimulation study three facilitatory ISIs were averaged separately, obtaining grandmean amplitudes of the inhibitory and of the facilitatory ISIs.
Magnetic stimulation was performed using two high-power Magstim The test after Rimonabant was performed using cortical mag- 200 magnetic stimulators (Magstim Co., Whitland, UK) connected to netic stimuli of the same intensity as the test and the conditioning the Bistim Module throughout all measurements. A figure-of-eight stimuli used in baseline conditions. Additionally, when there was a coil with external loop diameters of 9 cm was held over the right change in threshold or MEP amplitude, paired stimulation was motor cortex at the optimum scalp position to elicit motor responses repeated with the intensity of the cortical stimulus adjusted in order in the contralateral first dorsal interosseus (FDI). The induced to maintain a control MEP of the same amplitude as that recorded in current flowed in a postero-anterior direction. Resting motor baseline conditions and the conditioning stimulus intensity adjusted threshold (RMT) was defined as the minimum stimulus intensity in order to maintain an intensity of 90% of the AMT that produced a liminal motor evoked response (about 50 μV in 50% portant because the intensities of both conditioning and test stimuli of 10 trials) at rest. Active motor threshold (AMTM) was defined as have a profound effect on the degree of inhibition/facilitation the minimum stimulus intensity that produced a liminal motor produced by paired magnetic stimulation evoked response (about 200 μV in 50% of 10 trials) during isometric ). In studies involving drugs that affect motor contraction of the tested muscle at about 20% maximum. A constant thresholds, the interpretation of the results of TMS paired-pulse level of voluntary contraction was maintained with reference to an protocols which involve threshold-adjusted stimuli such as SICI and oscilloscope display of EMG in front of the subject. Auditory ICF might be problematic. One solution to the problem is to adjust feedback of the EMG activity was also provided.
the test stimulus intensity after drug in order to control the degree Motor evoked potentials (MEP) was obtained while subjects held of corticospinal tract activation ). This is relevant a tonic voluntary contraction of approximately 20% of maximum because it has been shown that in paired pulse protocols only the voluntary contraction (MVC) using a stimulus intensity of 120 and late I-waves are inhibited or facilitated ( 150% AMTM (in a total of ten trials). A constant level of voluntary Thus with a comparable degree of corticospinal tract activation a contraction was maintained with reference to an oscilloscope similar composition of the descending volleys will be obtained display of EMG in front of the subject. Auditory feedback of the before and after drug administration and any change in the amount EMG activity was also provided.
of inhibition or facilitation could be demonstrated. Ideally, to Cortical silent period (CSP) was elicited while subjects held a compare inhibition and facilitation a full intensity curve with tonic voluntary contraction of approximately 50% of MVC. Five multiple intensities fo the test and conditioning stimulus both stimuli at 120 and 150% AMTM were given. The duration of the CSP before and after the drug should be obtained. However, this makes was measured from the end of the motor potential to the resumption the procedure extremely time consuming and might interfere with (at any level) of sustained EMG activity. The test after Rimonabant full cooperation of the subjects. Thus, a single adjusted intensity was performed using cortical magnetic stimuli of the same intensity was evaluated in the present study. A MEP of 1 mV requires the as used in baseline conditions. Additionally, when there was a recruitment of several late I-waves the change in threshold or amplitude, TMS was repeated with the waves that are modulated in paired pulse protocols. For this reason, intensity of the cortical stimulus adjusted in order to maintain a this amplitude is particularly useful when evaluating paired pulse control MEP of the same amplitude as that recorded in baseline inhibtion/facilitation. A conditioning stimulus intensity of 90% AMT conditions. This is important because the MEP amplitude has a produces a strong cortical inhibition and facilitation without evoking profound effect on the CSP duration ( any descending activity (Thus, it appears In six subjects, we also performed electrical stimulation of the extremely useful in order to evaluate changes at cortical level. We motor cortex during voluntary contraction. This was performed will report the results obtained using both the same intensity of with a Digitimer D180A stimulator, with a time constant of 50 μs. The baseline conditions and the intensity of cortical stimulus adjusted.
cathode was located at the vertex and the anode 7 cm laterally On the other hand, only the data obtained using the intensity of (anodal stimulation). We evaluated electrical active motor thresh- cortical stimulus adjusted have a physiological interest, and – for old (AMTE) defined as the minimum stimulus intensity that produced this reason – only the adjusted data will be considered in the a liminal motor-evoked response of about 200 μV in 50% of 10 trials The same approach has been used also for long latency during voluntary contraction.
intracortical inhibition and short latency afferent inhibition (seebelow).
2.4. Paired pulse transcranial magnetic stimulation study Long latency intracortical inhibition (LICI) was studied as follows (). Using a Bistim Short latency intracortical inhibition (SICI) and intracortical facili- module, two magnetic stimuli were given through the same tation (ICF) were studied using the technique of .
stimulating coil over the motor cortex and the effect of the first Using a Bistim module, two magnetic stimuli were given through the (conditioning) stimulus on the second (test) stimulus was investi- same stimulating coil over the motor cortex and the effect of the gated. The conditioning and test stimulus was set at an intensity to first (conditioning) stimulus on the second (test) stimulus was in- evoke a muscle response in relaxed FDI with an amplitude of vestigated. The conditioning stimulus was set at an intensity of 90% approximately 1 mV, peak-to-peak. ISI of 100 ms was investigated. A of AMTM. The second, test, stimulus intensity was adjusted to evoke total of 15 conditioned and of 15 test responses were recorded. Each a muscle response in relaxed FDI with an amplitude of approximately paired-pulse TMS was pseudorandomly repeated and intermixed 1 mV, peak-to-peak. The timing of the conditioning stimulus was with the test stimuli. The amplitude of the conditioned MEPs was altered in relation to the test stimulus. Inhibitory interstimulus expressed as the percentage of the amplitude of the test EMG intervals (ISIs) between 2 and 5 ms in 1 s step (4 ISIs) and facilitatory responses. The test after Rimonabant was performed using cortical ISIs of 10, 15 and 25 ms were investigated. Five stimuli were de- magnetic stimuli of the same intensity as the test and the con- livered at each ISI. A total of 30 test responses were also recorded.
ditioning stimuli used in baseline conditions. Additionally, when Paired-pulse TMS was delivered at seven ISIs in the range of 2–25 ms there was a change in threshold or amplitude, paired stimulation was (specifically: seven conditions in which the conditioning stimulus repeated with the intensity of the cortical stimulus adjusted in order Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), doi: A. Oliviero et al.
to maintain a control MEP (both conditioning and test) of the same the test and the conditioning stimuli used in baseline conditions.
amplitude as that recorded in baseline conditions. We will report the Additionally, when there was a change in threshold or amplitude, results obtained using both the same intensity of baseline conditions paired stimulation was repeated with the intensity of the cortical and the intensity of cortical stimulus adjusted. On the other hand, stimulus adjusted in order to maintain a control EMG response of the only the data obtained using the intensity of cortical stimulus same amplitude as that recorded in baseline conditions. We will adjusted have a physiological interest, and – for this reason – only report the results obtained using both the same cortical stimulus the adjusted data will be considered in the intensity of baseline conditions and the intensity of cortical stimulus Short latency afferent inhibition (SAI) was studied using the adjusted. On the other hand, only the data obtained using the technique that has previously been described intensity of cortical stimulus adjusted have a physiological interest, Conditioning stimuli were single pulses and – for this reason – only the adjusted data will be considered in (200 μs) of electrical stimulation applied through bipolar electrodes to the median nerve at the wrist (cathode proximal). The intensity ofthe conditioning stimulus was set at just over motor threshold toevoke a visible twitch of the thenar muscles. The intensity of the 2.5. Statistical analysis cortical magnetic shock test was adjusted to evoke a muscleresponse in relaxed FDI with an amplitude of approximately 1 mV The effects of drug were tested separately for the TMS measures peak-to-peak. ISIs from 22 ms to 28 ms were investigated in 1 s step (RMT, AMT, MEP amplitude, SICI, ICF, LICI, SAI and CSP) and TES (7 ISIs). Five stimuli were delivered at each ISI and 30 test responses parameters (AMT and MEP amplitude) using ANOVA for repeated were obtained. For the conditioned responses, the conditioning measures, with the Greenhouse–Geisser correction for lack of peripheral nerve stimulus preceded the TMS test pulse at seven ISIs sphericity. Sphericity was tested using the Mauchly test. ANOVA in the range of 22–28 ms. Each interval was pseudorandomly significance was assumed whenever p b 0.05. We performed a large repeated and intermixed with the test stimuli (six conditions with number of ANOVAs and we did not apply corrections for multiple only the test stimulus repeated 5 times). The amplitude of the comparisons. This implies that the results are more sensitive than conditioned MEPs was expressed as a percentage of the amplitude of specific. We reported the exact p value so the reader can judge the test MEPs. The percentage of inhibition of the conditioned himself the robustness of each test. In the case of significant F responses at the seven different ISIs was averaged to obtain a grand values, post hoc analysis was performed using Tukey test.
mean (from a total of 35 trials). The test after Rimonabant was Significance was assumed whenever p b 0.05 (corrected for multiple performed using cortical magnetic stimuli of the same intensity as comparisons). Statistical tests concerning paired pulse experiments Baseline 2 Hours 24 Hours 48 Hours
Baseline 2 Hours 24 Hours 48 Hours
24 Hours 48 Hours
24 Hours 48 Hours
24 Hours 48 Hours
Time course of the TMS and TES motor thresholds and of MEP amplitudes obtained with fixed stimulus intensities of transcranial magnetic stimulation (TMS) and transcranial electrical stimulation (TES) before and after 20 mg intake of Rimonabant.
A. Resting motor threshold (RMT) and active motor threshold (AMT), tested using TMS are decreased by the drug intake for up to24–48 h. B. AMT, tested using TES, is decreased by the drug intake for up to 24 h. C. MEP amplitudes, tested using TMS at rest, areincreased by the drug intake. D. MEP amplitudes, tested using TMS during voluntary contraction, are increased by the drug intake whenthe intensity used was 120%AMT (gray dotted line). When the intensity used was 150%AMT, MEP amplitudes are unchanged by the drugintake (gray line). E. MEP amplitudes, tested using TES during voluntary contraction, are increased by the drug intake. (*P b 0.05).
Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), Effects of a CB1 antagonist on cortical excitability were performed on conditioned MEP amplitude values expressed as a significantly increased the mean MEP amplitude at rest. Post percentage of the test response.
hoc analysis showed that the effects of Rimonabant on MEPamplitude at rest were significant 24 h after the drug intakeThe effect was not significant anymore 48 h later.
Administration of Rimonabant significantly increased the meanMEP amplitude during slight voluntary contraction of the target Neurophysiological results are summarized in .
muscle when TMS intensity was set at 120% AMT. Post hoc Statistical analysis is reported in details in the table.
analysis showed that the effects of Rimonabant on MEPamplitude at 120% AMT were significant 24 h after the drug 3.1. Single pulse studies intake (D). The effect was not significant anymore 48 hlater. MEP amplitude was not significantly modified when 3.1.1. Transcranial magnetic stimulation tested at rest or during activation when a TMS intensity of 150%AMT was used. No effects were observed on CSP at both 120 and Administration of Rimonabant significantly reduced the meanAMT m, while RMT was less affected Post hoc analyses revealed that the effects of Rimonabant on AMTmstarted immediately (2 h) after the intake of the drug and 3.1.2. Transcranial electrical stimulation lasted up to 48 h, while the effects on RMT were significant only Administration of Rimonabant significantly reduced the mean 24 h after the drug intake. Administration of Rimonabant AMTe (B, ). Post hoc analysis showed that the % of test response 20 % of test response same intensity of baseline intensity of the cortical same intensity of baseline intensity of the cortical stimulus adjusted stimulus adjusted % of test response % of test response same intensity of baseline intensity of the cortical same intensity of baseline intensity of the cortical stimulus adjusted stimulus adjusted same intensity of baseline intensity of the cortical same intensity of baseline intensity of the cortical stimulus adjusted stimulus adjusted Time course of the short latency intracortical inhibition (SICI), intracortical facilitation (ICF), long latency intracortical inhibition (LICI), short latency afferent inhibition (SAI) and cortical silent period (CSP) before and after 20 mg intake of Rimonabant.
The test after Rimonabant was performed using cortical magnetic stimuli of the same intensity as the test and the conditioning stimuliused in baseline conditions (left bars of each graph). Additionally, when there was a change in threshold or amplitude, pairedstimulation was repeated with the intensity of the cortical stimulus adjusted in order to maintain a control EMG response of the sameamplitude as that recorded in baseline conditions (right bars of each graph). SICI was significantly reduced after drug intake, but whenthe test was performed using intensity of the cortical stimulus adjusted, SICI resulted unchanged. When the test was performed usingintensity of the cortical stimulus adjusted, ICF was significantly increased after drug intake. No changes are produced by theRimonabant on LICI, SAI and CSP. Histograms are mean values and error bars are standard deviations (*P b 0.05).
Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), doi: A. Oliviero et al.
Statistical analysis (ANOVA for repeated measures).
Main effect for TIME Baseline vs Post 1 Baseline vs Post 2 Baseline vs Post 3 MEP amplitudeRest (TMS) Active 120% AMT (TMS) 8.725 [1.3,10.7] 0.0004 0.1079 Active 150% AMT (TMS) Active 120% AMTe (TES) Cortical silent periodsActive 120% AMT Active 120% AMT (MEP amplitude matched) Active 150% AMT (MEP amplitude matched) Paired pulse studiesSICI (2–5 ms ISIs) ICF (10–15–25 ms ISIs) SAI (2–5 ms ISIs) LICI (100 ms ISI) SICI (2–5 ms ISIs) (matched) ICF (10–15–25 ms ISIs) (matched) SAI (2–5 ms ISIs) (matched) LICI (100 ms ISI) (matched) effects of Rimonabant on AMTe started immediately (2 h) after plitude was increased by Rimonabant, we repeated the the intake of the drug and lasted at least 24 h. The effect was experiments after adjusting the stimulus intensities to com- not significant anymore 48 h later. Administration of Rimona- pensate for the changes in threshold and EMG response bant significantly increased the mean MEP amplitude during amplitude after Rimonabant. With adjusted stimulus intensi- slight voluntary contraction of the target muscle. Post hoc ties, the amount of facilitation was significantly increased 24 h analysis showed that the effects of Rimonabant on MEP after the oral dose of Rimonabant.
amplitude at 120% AMT started immediately (2 h) after theintake of the drug and lasted at least 24 h E). The effect 3.2.2. Long latency intracortical inhibition was not significant anymore 48 h later.
When two stimuli were delivered, the EMG responses to the teststimulus were dramatically suppressed at 100 ms ISI. The 3.2. Paired pulse studies amount of inhibition was unchanged by the administration of20 mg of Rimonabant (Because AMT was decreased and 3.2.1. Short latency intracortical inhibition and EMG response amplitude was increased by Rimonabant, we repeated the experiments after adjusting the stimulus inten- When two stimuli were delivered, the EMG responses to the test sities to compensate for the changes in threshold and EMG stimulus were dramatically suppressed at ISIs of 2–5 ms ).
response amplitude after Rimonabant. With adjusted stimulus The amount of inhibition over ISIs of 2–5 ms was reduced by the intensities and EMG response amplitudes, the amount of administration of 20 mg of Rimonabant. Because AMT was inhibition was not modified by Rimonabant.
decreased and EMG response amplitude was increased byRimonabant, we repeated the experiments after adjusting the 3.2.3. Short latency afferent inhibition stimulus intensities to compensate for the changes in threshold When two stimuli were delivered (one TMS stimulus preceded and EMG response amplitude after Rimonabant. With adjusted by a peripheral electrical stimulus of the median nerve), the stimulus intensities and EMG response amplitudes, the amount EMG responses to the test stimulus were dramatically sup- of inhibition over ISIs of 2–5 ms was not modified by pressed at ISIs of 22–28 ms. The amount of inhibition was unchanged by the administration of 20 mg of Rimonabant When two stimuli were delivered, the EMG responses to the (). Because AMT was decreased and EMG response ampli- test stimulus were increased at ISIs of 10–25 ms. The amount tude was increased by Rimonabant, we repeated the experi- of facilitation was not significantly modified by Rimonabant ments after adjusting the stimulus intensities to compensate for ). Because AMT was decreased and EMG response am- the changes in threshold and EMG response amplitude after Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), Effects of a CB1 antagonist on cortical excitability Rimonabant. With adjusted stimulus intensities and EMG in humans as tested by transcranial magnetic and electric response amplitudes, the amount of inhibition was not modified stimulations. These results are in line with experimental by Rimonabant.
data reporting that Delta 9-THC increases the firingthreshold for the motor neuron action potential in cats () and that Rimonabant(SR141716A) enhances spinal reflexes in rabbits The present results provide the first evidence that the Another important observation is that Rimonabant seems excitability of motor corticospinal networks as tested by to increase spinal cord excitability in a state dependent transcranial magnetic and electrical stimulation in humans manner. Voluntary contraction – which normally reduces can be increased by a single oral dose of 20 mg of the CB1 TMS thresholds and increases MEP amplitudes – magnifies the receptor antagonist/inverse agonist Rimonabant.
observation of the extra facilitation produced by Rimona-bant. In fact, if the spinal changes would be present also at 4.1. Responses to single magnetic and electrical rest with a similar amount, we would expect a more intense reduction of the RMT and a stronger increase of MEP am-plitudes at rest. Moreover, both thresholds and MEP TES directly activates the corticospinal axons, and for this amplitudes are more affected during voluntary contraction.
reason it is a useful test of the change of excitability of the This is in agreement with the location of CB1 on presynaptic central motor pathways at spinal cord level ( axons and with its role in controlling transmitter release ). More in details, AMTE reflects the intrinsic and extrinsically modulated excitability properties of spinal ). Endocannabinoid release can be triggered by motor neurons and of corticospinal axons at a distance from calcium influx during large depolarizations and in this way it the soma and the axon hillock of the cortical motor neurons would control the transmitter release. This mechanism could be under the well-known effect of cannabinoids in spasticity, activates corticospinal pathways trans-synaptically, so it is reducing spinal motor neurons hyperexcitability useful to study motor cortex excitability. Further, RMT and ). In our experiment AMTM reflect the intrinsic and extrinsically modulated motor neuron depolarization is activated by voluntary excitability properties of corticospinal neurons ( contraction and the endocannabinoids modulation is pre- vented by CB1R blockade. Spinal motor neurons have a small Rimonabant reduced active and resting motor thresholds.
amount of tonic discharge and for this reason the effects at The reduction in motor threshold was accompanied by an rest seems to be much smaller.
increase in the amplitude of EMG responses evoked by mag-netic stimulation at rest and during voluntary contractionwith the lower intensity studied (120% AMT 4.2. Paired pulse studies particularly evident 24 h after the drug intake. Theamplitude of EMG responses evoked by magnetic stimulation Short latency intracortical inhibition (SICI) to paired pulse during voluntary contraction at the highest intensity studied TMS, long latency intracortical inhibition (LICI) to paired (150% AMTM) did not show a significant increase. This lack of pulse TMS and the cortical silent period (CSP), are all be- increase could be due to a "ceiling" effect, or it might be lieved to reflect the excitability of inhibitory GABAergic also due to the recruitment of different populations of cortical circuits. In addition, SICI reflects the activity of interneurons or motor neurons depending on the intensity of GABAergic inhibitory circuits mediated by GABAA receptors.
voluntary contraction, that might respond differently to However, LICI reflects the activity of GABAergic inhibitory circuits mediated by GABAB receptors. The mechanisms Similarly to the MEPs evoked by TMS, the responses generating CSP are – at least in part – different from those evoked by electrical stimulation during voluntary contrac- responsible for SICI, in that CSP appears to be more related tion were facilitated by administration of Rimonabant. Thus, to GABAB mechanisms However, the electrically evoked responses are as sensitive to changes in administration of lorazepam – a benzodiazepine which excitability as those evoked by magnetic stimulation. We determines an increase of chloride channel opening fre- interpret that a similar effect of Rimonabant on EMG re- quency through the activation of GABAA receptors – induces sponses evoked by magnetic and electric stimulations a lengthening of CSP duration in normal subjects indicates that the increase in excitability takes place – ). It is therefore conceivable that, at least in perhaps not exclusively (see below) – at the level of circuits part, silent period also depends on GABAA activity, and the within the spinal cord. Latency analysis of MEPs evoked by CSP could be considered a mixed effect of activation of both TMS also confirmed a role of spinal cord circuits in hyper- GABAA and GABAB receptors.
excitability induced by Rimonabant. Latencies were reduced A single oral dose of 20 mg of Rimonabant had no effects of about 1–1.6 ms. This corresponds to the normal interpeak on all the inhibition that are thought to be GABAergic in latency of I waves evoked by TMS origin. This is apparently in contrast with the reduction The reduced latency means that spinal motor neurons are of GABAergic (GABAA) inhibition reported in chronic canna- discharging when an earlier I wave is arriving. This could be bis consumers (Rimonabant is an due to a more excitable spinal motor neuron. The present antagonist/inverse agonist of CB1 receptors, therefore we results provide the first evidence that blockade of CB1re- could expect an opposite effect with respect to the cannabis ceptors can increase the excitability of spinal motor circuits users — e.g., increased GABAA inhibition with the effects Please cite this article as: Oliviero, A., et al., CB1 receptor antagonism/inverse agonism increases motor system excitability in humans,Eur. Neuropsychopharmacol. (2011), doi: A. Oliviero et al.
detectable by increased SICI and prolonged CSP, but this was protocols with authors A. Arevalo-Martin, D. García-Ovejero, L.
not the case. Differences in chronic vs acute intake of the Chiovato, J. Aguilar, G. Foffani, and V. Di Lazzaro.
drug, or differential effects over different GABAergic Authors L. Mordillo-Mateos, A. Lozano-Sicilia, and I. Panyavin per- populations of interneurons may account for the differential formed the experiments and the initial data processing and analysis.
Authors A. Oliviero, M. Rotondi, J. Aguilar, and G. Foffani, effects on SICI. Indeed, CB1 can be expressed either in performed the statistical analyses and wrote the first draft of the excitatory or inhibitory cells of cortical and subcortical paper. All authors contributed to and have approved the final structures and both excitatory and inhibitory effects have been described in different experi-mental systems depending on the dose and context ofcannabinoid agonist or antagonist treatments ( Conflict of interests The authors (AO, AAM, MR, DGO, LMM, ALS, LC, JA, GF, EMH) declare Intracortical facilitation (ICF) to paired pulse TMS is that part of the data presented here has been used for the redaction thought to depend upon the activity of intracortical of PCT/ES2009/000010 and PCT/ES2010/070012 (patent pending).
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dependent manner how Rimonabant facilitate motor neurons Di Lazzaro, V., Oliviero, A., Profice, P., Saturno, E., Pilato, F., at spinal level. The conditioning stimulus lowers the Insola, A., Mazzone, P., Tonali, P., Rothwell, J.C., 1998a.
corticospinal cell threshold to fire when non NMDA glutama- Comparison of descending volleys evoked by transcranial tergic neurotransmission is facilitated (Di Lazzaro et al., magnetic and electric stimulation in conscious humans. Ele- 2003). Again, this is in agreement with the location of CB1 on croencephalography and Clinical Neurophysiology 109, 397–401.
presynaptic axons and with its role in controlling transmitter Di Lazzaro, V., Restuccia, D., Oliviero, A., Profice, P., Ferrara, L., Insola, A., Mazzone, P., Tonali, P., Rothwell, J.C., 1998b. Effects Conditioning stimulus can activate of voluntary contraction on descending volleys evoked by glutamate release at cortical synapses and due to the fact transcranial stimulation in conscious humans. The Journal of that the endocannabinoids modulation is prevented by CB1R Physiology 508, 625–633.
Di Lazzaro, V., Oliviero, A., Profice, P., Insola, A., Mazzone, P., blockade the test stimulus is more effective.
Tonali, P., Rothwell, J.C., 1999a. Effects of voluntary contrac- In conclusion, our results demonstrate that a single oral tion on descending volleys evoked by transcranial electrical dose of 20 mg of Rimonabant can increase motor system stimulation over the motor cortex hand area in conscious excitability at cortical and spinal levels. This opens new humans. Experimental Brain Research 124, 525–528.
avenues to test the CB1R antagonists/inverse agonists for the Di Lazzaro, V., Rothwell, J.C., Oliviero, A., Profice, P., Insola, A., treatment of neurological dysfunctions in which can be useful Mazzone, P., Tonali, P., 1999b. Intracortical origin of the short to increase the excitability levels of motor system. Virtually latency facilitation produced by pairs of threshold magnetic stimuli all the disorders characterized by a reduced output of the applied to human motor cortex. Experimental Brain Research 129, motor cortex can be included in the list of the disorders that can be treated using CB1 antagonists/reverse agonists (e.g.
Di Lazzaro, V., Oliviero, A., Mazzone, P., Pilato, F., Saturno, E., Insola, A., Visocchi, M., Colosimo, C., Tonali, P.A., Rothwell, stroke, traumatic brain injury, spinal cord injury, multiple J.C., 2002. Direct demonstration of long latency cortico-cortical sclerosis, fatigue syndromes, parkinsonisms, etc.).
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Role of the funding source Di Lazzaro, V., Oliviero, A., Profice, P., Pennisi, M.A., Pilato, F., Zito, G., Dileone, M., Nicoletti, R., Pasqualetti, P., Tonali, P.A.,2003. Ketamine increases human motor cortex excitability to This study was funded by the Fundación Hospital Nacional de transcranial magnetic stimulation. The Journal of Physiology 547 Parapléjicos. The Fundación Hospital Nacional de Parapléjicos had (Pt 2), 485–496.
no further role in study design, in the collection, analysis and Di Lazzaro, V., Oliviero, A., Pilato, F., Saturno, E., Dileone, M., interpretation of data, in the writing of the report and in the Mazzone, P., Insola, A., Tonali, P.A., Rothwell, J.C., 2004. The decision to submit the paper for publication.
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