APPARATUS AND METHOD FOR TREATING A CORTICAL-BASED VISUAL DISORDER USING TRANSCRANIAL MAGNETIC STIMULATION

Field The present invention relates to the field of transcranial magnetic stimulation. The present invention also relates to the field of treatment of cortical-based visual disorders.

Background Amblyopia, a cortically based visual disorder caused by disruption of visual development during an early developmental critical period, is thought to be a largely intractable problem in adult patients due to a lack of plasticity after this critical period. Monocular amblyopia is the largest cause of uniocular impairment in the adult population with an incidence of 3%. Current treatment approaches emphasize patching or penalization of the non-amblyopic eye before 12 years of age. There is no treatment available for individuals outside of this critical period and although new approaches are being explored, current treatment alternatives are limited and often unsuccessful.

Amblyopia is a neurological disorder with a cortical basis, and no treatment approaches to date have aimed to directly affect the neural processing in visual cortex, rather either general pharmacological interventions have been used or treatments requiring the patching or penalization of one eye.

Summary of the Invention

The applicants have invented a process and apparatus for using repetitive transcranial magnetic stimulation (rTMS) to improve vision in patients suffering from cortical-based visual disorders such as amblyopia. rTMS is a way of directly influencing the neural processing of a fairly specific cortical region by repeatedly administering magnetic pulses to the head through a specially designed coil. The magnetic field sets up an electrical current in the cortical tissue underneath it causing a population of the neurons in that tissue to fire. When applied in a repetitive manner TMS can have effects on cortical regions

that last longer than the rTMS session itself. Transcranial magnetic stimulation has also been studied as a potential therapy for other diseases such as stroke, aphasia, Parkinson's disease, amyotrophic lateral sclerosis, epilepsy, migraine.

The applicants have discovered that there is a therapeutic effect in amblyopic patients. The rTMS administration includes a range of administration frequencies (number of TMS pulses per second) and durations (the length of time for which the rTMS is applied). In addition, the invention includes the apparatus required for the correct administration of the rTMS, including methods and apparatus for selecting the correct stimulation site in visual cortex which varies between patients, the correct stimulation intensity to use which also varies and criteria for the exclusion of patients for whom rTMS treatment is not appropriate.

Applicants present data showing that repetitive transcranial magnetic stimulation (rTMS) of the visual cortex can temporarily reverse what has been considered an irreversible visual loss in adult amblyopes. The results indicate continued plasticity of the brain in adulthood and open the way for new therapeutic approach to the treatment of this condition. Applicants hypothesize that the effects of rTMS are mediated by changes in intra-cortical inhibition (ICI) within the visual cortex and therefore may directly link recent physiological studies of amblyopia recovery in animals with improvement in human patients. It has been shown that Environmental enrichment in adulthood promotes amblyopia recovery through a reduction of intracortical inhibition in rats (Sale, A., Maya Vetencourt, J. F., Medini, P., Cenni, M. C, Baroncelli, L., De Pasquale, R., and Maffei, L. 2007. Nat Neurosci 10, 679- 681 ). Applicants use contrast sensitivity as the measure of therapeutic benefit in their experiments. Contrast sensitivity is defined as the lowest contrast at which a person is able to reliably see a visual stimulus with a fixed spatial frequency. Spatial frequency is related to the size of the features of the stimulus. A grating with wide bars/stripes has a low spatial frequency as the pattern repeats itself infrequently (because the bars are wide). A grating with

very thin bars has a high spatial frequency as the pattern repeats itself frequently. Spatial frequency is measured in cycles per degree, the number of repetitions of the pattern within a single degree of visual angle. The higher the cycles per degree, the higher the spatial frequency.

It is an object of the present invention to provide a treatment control system for treating a cortical-based visual disorder using delivery of repetitive transcranial magnetic stimulation (TMS) to the visual cortex of a patient, the system comprising one or more of a module for analyzing a patient's brain imaging data to generate coil position data for setting a position of a TMS coil, a module for recording a position of phosphenes within a field of view experienced by the patient to generate coil position data for adjusting and/or setting a position of a TMS coil and a module for recording the experience of phosphenes by the patient for a number of TMS events to determine parameters for repetitive TMS therapy. The system could further comprise an interface for interfacing said coil position data with a robot for positioning said TMS coil. The system could further comprise a module for physically positioning, repositioning and fixing a stimulation coil over a head of the patient. The system could further comprise a module for recording vision test results from a patient before and after delivery of said stimulation to determine improvement of vision as a result of said stimulation. The system could further comprise a user input device which allows the patient to enter responses during a vision task or during the phosphene experience phase for determining optimal location, intensity and frequency of stimulation.

It is an object of the present invention to provide an apparatus for treating a cortical-based visual disorder using delivery of repetitive transcranial magnetic stimuli to the visual cortex of a patient comprising a stimulation coil for delivering transcranial magnetic stimulation, said coil positioning adjusted over visual cortex on the basis of patient responses to the location of phosphenes in the visual field; and stimulation intensity adjusted on the basis patient comfort; and a control system which analyzes patient responses and adjusts any one of or any combination of stimulation location, frequency and intensity for maximal

therapeutic benefit to said patient. This apparatus can comprise a module for recording vision test results from a patient before and after delivery of said stimulation to determine improvement of vision as a result of said stimulation.

It is a further object of the present invention to provide a treatment control system for treating a cortical-based visual disorder such as amblyopia using delivery of repetitive transcranial magnetic stimulation to the visual cortex of a patient, the system comprising a module for recording vision test results from a patient before and after delivery of said stimulation to determine improvement of vision as a result of said stimulation; and a module for recording phosphenes experienced by the patient to determine an intensity of said stimulation and/or a position of said stimulation. Although the stimulation coil can be positioned manually by an operator, the system also comprises a module/robotic arm for physically positioning, repositioning and fixing the stimulation coil over the head of a patient automatically, based on gross anatomical landmarks.

It is yet another object of the present invention to provide an apparatus for treating a cortical-based visual disorder such as amblyopia using delivery of repetitive transcranial magnetic stimuli to the visual cortex of a patient comprising a stimulation coil for delivering transcranial magnetic stimulation at a fixed location over the visual cortex, at a fixed intensity to ensure safety and a fixed frequency that does not induce phosphenes or discomfort in substantially all patients tested. This apparatus can comprise a device such as a robotic arm which is interfaced with the treatment control system and designed to automatically position the stimulation coil over the visual cortex on the basis of gross anatomical structures of the head. Alternatively, the coil can be integrated into a cap-like device which automatically espouses the shape of the patient's head. If the patient to be treated has previously been the object of any type of medical imaging event such as computerized tomography (CT) or magnetic resonance imaging (MRI), the image data arising from these events can be utilized to identify the specific location of the visual cortex for subsequent delivery of transcranial magnetic stimulation. This device can be

adapted for treatment of a patient outside a clinical or hospital setting such as in the home of a patient.

It is an object of the present invention to provide a method for the treatment of visual disorders wherein the stimulation frequency is set at the lowest value at which practically all patients respond. Applicant's experiments have shown that optimal frequency of stimulation is greater than 1 Hz as only a portion of patients responded at this frequency. Because 10 Hz was shown by applicants to increase contrast sensitivity in all patients tested, the optimal frequency is approximately equal to 10Hz.

In another embodiment of the present invention, the treatment control system comprises a user input device such as a dial, a button or any structure that allows a patient to select responses in a visual discrimination task for contrast sensitivity such as, but not limited to, letter-based, motion or orientation discrimination. The purpose of these visual tasks, which can be performed monocularly or binocularly are to allow the system or the operator to evaluate the effect of transcranial magnetic stimulation. If the treatment control system or the operator determines an improvement in contrast sensitivity, then the stimulation location and intensity of the coil can be maintained. If however the visual task does not allow the system or operator to determine an improvement in some measure of visual acuity, then the location of stimulation can be changed and the intensity of stimulation can be increased.

It is an object of the present invention to provide a apparatus which combines binocular vision therapy, a new technique also developed by applicants and described in the following publication (Mansouri B, Thompson B, Hess RF. 2008. Vision Research (48)28: 2775-2784. Measurement of suprathreshold binocular interactions in amblyopia) with an apparatus as described herein to deliver transcranial magnetic stimulation. Indeed, delivery of TMS immediately preceding, during or immediately after binocular training is thought to induce a synergistic therapeutic benefit to the patient which is greater than the sum of both therapies individually.

It is an object of the present invention to provide a method for treating a cortical-based visual disorder comprising delivery of transcranial magnetic stimulation to the visual cortex of a patient. This method can comprise testing visual function prior to said stimulation and testing visual function after said stimulation to determine improvement, wherein treatment is repeated as a function of said improvement. Visual function can be evaluated by any test of visual acuity and recorded either manually by an operator or automatically by the system itself.

It is yet another object of the present invention to provide a method wherein the stimulation frequency is set at the lowest frequency at which practically all patients respond. This allows the operator or system to easily set the initial frequency of stimulation. It has been shown by applicants that 1 Hz is beneficial only to a fraction of the patients and 10 Hz stimulation was beneficial to all patients tested. This suggests that the optimal frequency is between 1 and 10 Hz.

It is yet another object of the present invention to provide a method wherein stimulation location is automatically adjusted over the visual cortex so as to insure optimal position of phosphenes which are seen by the patient. It is important to centrally position the phosphenes in the visual field so that the coil is centrally positioned over the visual cortex, thus insuring that small variations in coil location will continue to stimulate the visual cortex.

Brief Description of the Drawings

Figure 1 shows the effects of 1 Hz rTMS on contrast sensitivity.

Figure 2 shows the effect of 1 Hz (a and c) and 10 Hz (b and d) rTMS over visual cortex for two patients who showed an increase in contrast sensitivity after 1 Hz rTMS for the amblyopic eye, high spatial frequency condition

Figure 3 is a schematic block diagram of the units embedded in the invention, namely the VISION-TMS package,

Figure 4 is a flow chart of the steps involved in one embodiment of the invention whereby repetitive TMS is applied to the primary visual cortex to treat a specific cortically based visual disorder, namely monocular amblyopia.

Figure 5 is a flow chart of the steps involved in a test for comfort for participants in one embodiment of the invention.

Figure 6 is a flow chart of the steps involved in screening for phosphenes (finding the optimal stimulation site) in one embodiment of the invention.

Figure 7 is a flow chart of the steps involved in finding the correct stimulation intensity at which to administer the rTMS.

Figure 8 is a complement to Figure 1 and comprises additional subjects demonstrating the effects of 1 Hz rTMS over visual cortex on contrast detection for amblyopic participants.

Figure 9 is a complement to Figure 2 and comprises additional subjects demonstrating the effects of 10 Hz rTMS over visual cortex on contrast sensitivity for amblyopic participants

Figure 10 is a 10-20 system representation of the skull and brain area of a patient.

Description of Embodiments

Brain plasticity has however been reported in adult visual cortex both in normal populations amblyopic humans and animals (Sale et al., 2007) suggesting that amblyopic visual cortex does possess some capacity for improved function. Recovery of visual function in post-critical period amblyopic animals has recently been suggested to be mediated in part by reduced ICI in visual cortex.

rTMS is a technique that has been shown to depress intracortical inhibition after long trains of subthreshold repetitive magnetic stimuli at low frequency in the human motor cortex (Modugno, N., Curra, A., Conte, A., Inghilleri, M., Fofi, L., Agostino, R., Manfredi, M., and Berardelli, A. 2003. Clin. Neurophysiol. 114, 2416-2422). This led to the hypothesis that rTMS over the visual cortex would also reduce ICI, thus improving vision in the amblyopic eye. rTMS has also been shown to modulate levels of brain derived neurotrophic factor (BDNF), a protein thought to be involved in recovery from monocular deprivation (Sale et al., 2007). There is currently no consensus on which stimulation regime maximally influences ICI so applicants initially used 1 -Hz low frequency rTMS due to its higher level of safety. Applicants initially chose to apply high frequency (10-Hz) rTMS only if patients did not respond to the low frequency stimulation and then added additional patients when the 10Hz rTMS had beneficial effects.

To quantify any effects of rTMS on amblyopic vision, applicants measured contrast sensitivity to one low (1 cycle per degree or cpd) and one high (either 10 or 20 cpd) spatial frequency in seven amblyopic observers directly before (t1 ), directly after (t2) and 30 minutes after (t3) 10 minutes of 1 Hz rTMS stimulation of the visual cortex shown in Figure 1. Contrast sensitivity was measured using a 2-up 1 down staircase technique with three measurements taken for each participant per eye/spatial frequency combination within each block of measurements (t1 -3). This method of measurement was used as it provided the best tradeoff between accuracy and speed of measurement as necessitated by the transient nature of rTMS effects. In order to control for non-rTMS related changes in contrast sensitivity a number of controls were built into the applicant's study. Figure 1 shows the effects of 1 Hz rTMS on contrast sensitivity. Measurements were made pre-rTMS (t1 ), directly post rTMS (t2) and 30 minutes post rTMS (t3) for both the amblyopic eye (AME) and the fellow fixing eye (FFE) for high spatial frequencies (a and c) (10cpd for 4 patients, 20 cpd for 1 patient) and for low spatial frequencies (b) (1 cpd). rTMS was administered over visual cortex (a and b) and motor cortex (c). rTMS over visual cortex significantly reduced contrast thresholds for the

amblyopic eye high spatial frequency condition only (Tukey's HSD test showed a significant improvement from t1 to t2 q(106.4)=5.6, p<.05 and a greater improvement from t1 to t3 q(106.4)=8.12, p<.01 ).

Firstly applicants tested high and low spatial frequency contrast sensitivity in both the amblyopic and the non-amblyopic (fellow fixing) eye of patients in all experimental sessions. This provided a measure of the effect of rTMS on normally functioning components of the visual system. Fellow fixing eyes do not show pronounced contrast sensitivity deficits. In addition amblyopic eyes typically do not show a pronounced deficit at low spatial frequencies. Therefore applicants had one experimental condition (amblyopic eye, high spatial frequency) and three control conditions where no improvement was anticipated (amblyopic eye, low spatial frequency, low and high spatial frequencies fellow fixing eye). In addition applicants ran a control experimental condition whereby rTMS was delivered over motor cortex. In this condition the patients experienced all the peripheral effects of TMS including a TMS induced effect, in this case a twitch in the left FDI muscle, but with no intra-cortical inhibition changes in visual cortex. Visual cortex rTMS was delivered over an optimal phosphene location close to the occipital poles. The optimal phosphene location was independently identified in each patient.

The applicant's results showed two clearly separate groups of patients. Five out of seven patients showed a significant increase in contrast sensitivity for the amblyopic eye, high spatial frequency condition after low frequency rTMS as can be seen in Figure 1A. Fellow fixing eyes, seen in Figure 1 B, and the amblyopic eye low spatial frequency sensitivities did not change. As shown in Figure 1 C, no changes were found for motor cortex stimulation. Figure 2 shows the effect of 1 Hz (a and c) and 10 Hz (b and d) rTMS over visual cortex for two patients who showed an increase in contrast sensitivity after 1 Hz rTMS for the amblyopic eye, high spatial frequency condition (a). Rerunning these participants with 10 Hz rTMS had a beneficial effect on contrast sensitivity for the amblyopic eye, high spatial frequency condition only (b).For the two remaining participants presented in Figure 2A, there was a clear decrease in

contrast sensitivity for the amblyopic eye high spatial frequency condition only. Following their protocol, applicants retested the two patients who showed a temporary decrease in contrast sensitivity using high-frequency (10 Hz) rTMS over visual cortex. This manipulation produced an improvement in both patients, as can be observed in Figure 2B.

Figure 3 is a schematic block diagram of the units embedded in the invention, namely the VISION-TMS package, which is applied for use of Transcranial Magnetic Stimulation (TMS) in the treatment of visual deficiencies with a cortical origin such as amblyopia. The accompanying apparatus to the TMS machine consists of (a) TMS coil positional control unit (b) central processing unit (c) patient behavioral response unit and (d) visual presentation unit.

a. TMS coil positional control unit: i. TMS cap with all the testing regions marked on it ii. Coil mounted on a helmet iii. 3D positioning system (with commercially available package brain-sight and MRI images) iv. Suspended non-automatic or semiautomatic weight bearing or robotic arm

b. Central processing unit: i. Microchips or software to control TMS positioning, intensity calibration and treatment administration. ii. TMS machine iii. Registration and treatment procedure iv. Receive the feedback from patient and analyze and react appropriately

c. Patient behavioral response unit: i. Eye-tracker ii. Keypad iii. Touch sensitive display

d. Visual presentation unit i. LCD goggles or optic fiber goggles ii. Regular monitors iii. Specialized display with touch sensitive screen

Figure 4 is a flow chart of the steps involved in one embodiment of the invention whereby repetitive TMS is applied to the primary visual cortex to treat a specific cortically based visual disorder, namely monocular amblyopia.

Figure 5 is a flow chart of the steps involved in a test for comfort for participants in one embodiment of the invention. Comfort testing is performed to define the maximum stimulation intensity that can be used without causing the patient discomfort.

Figure 6 is a flow chart of the steps involved in screening for phosphenes (finding the optimal stimulation site) in one embodiment of the invention. The algorithm that receives feedback from the patient regarding the strength and location of phosphenes calculates the stimulation site that provided the most reliable, strong phosphene in a central region of the visual field. Phosphene perception is also verified by the algorithm by ensuring that phosphenes are distributed in a retinotopic nature around the visual field.

Figure 7 is a flow chart of the steps involved in finding the correct stimulation intensity at which to administer the rTMS. Stimulation calibration is achieved using reported phosphenes from the participant.

Figure 8 is a complement to Figure 1 and comprises additional subjects demonstrating the effects of 1 Hz rTMS over visual cortex on contrast detection for amblyopic participants. For both the amblyopic eye and the fellow fixing eye, measurements were made before rTMS (TO), directly after rTMS (T1 ), and 30 min after rTMS (T2) for a high spatial frequency and for a low spatial frequency. Data for T1 and T2 were normalized to the baseline (T0-T1

and T0-T2) and plotted on the y axis as a change in percentage of contrast relative to TO. A positive difference therefore indicates an improvement in contrast sensitivity (more contrast required before rTMS than after). Error bars represent ± 1 standard error of the mean (SEM); n = 9.

Figure 9 is a complement to Figure 2 and comprises additional subjects demonstrating the effects of 10 Hz rTMS over visual cortex on contrast sensitivity for amblyopic participants grouped in Figure 9A and individual data in Figures 9B and 9C and presented as in Figure 1 (n = 6), for the amblyopic eye, high spatial frequency condition.

Figure 10 is a 10-20 system representation of the skull and brain area of a patient. The 10-20 system is an internationally known method to describe the location of scalp areas thus helping in the positioning of electrodes on said scalp for electroencephalography (EEG) procedures. The system is based on the relationship between the location of an electrode and the underlying area of cerebral cortex wherein 10 and 20 refer to the percentage difference in distances between adjacent electrodes with respect to the front-back or right- left distance of the skull. The primary visual cortex (V) is found mainly in the occipital lobe which is described as O1 and O2 on the drawing.

rTMS over visual cortex was found to have a beneficial effect in the applicant's sample of amblyopic patients. Contrast sensitivity was improved for high spatial frequencies in the amblyopic eye directly after rTMS and 30 after rTMS. Applicants hypothesize that this effect is mediated by reduced intra-cortical inhibition in visual cortex after rTMS which facilitates improved visual performance. There are, however, other possible mechanisms of rTMS action that should be considered. Low and high speed rTMS are thought to decrease and increase excitability of the stimulated region respectively. The efficacy of rTMS has been shown to rely on the recent history of activity in the stimulated neurons (Iyer, M. B., Schleper, N., and Wassermann, E. M., 2003. J Neurosci 23, 10867-10872). Therefore, if neurons activated by input from the amblyopic eye differ in their activation level relative to fellow fixing eye neurons, then

rTMS will differentially affect these two neuronal populations. Depending on the sign of the difference in historical activity, either high or low frequency rTMS could be required to match or shift the balance in neuronal excitability in favor of the amblyopic eye and therefore increase acuity, either through a removal of inhibition of inputs from the amblyopic eye by the fellow eye or through facilitating the activity of neurons driven by the amblyopic eye.

A related consideration is the effect of rTMS on the neurochemistry of the stimulated region. It has been shown that increasing the global levels of dopamine in the brain improves acuity in the amblyopic eye and rTMS over motor cortex has been shown to increase dopamine release. However given the paucity of dopaminergic innervation in the visual cortex increase in local dopamine release may not in itself be a sufficient explanation for the effects of rTMS shown here. Furthermore, the hypothesis that a global level of dopamine increase may be responsible for improved acuity after rTMS does not account for the lack of an effect of rTMS over motor cortex on visual acuity, which presumably would also influence dopamine levels. Resolution of these issues will require further investigation in physiological and neuroimaging domains. The current data, obtained in a small sample of amblyopic persons, suggest that rTMS may be a useful tool for the investigation and treatment of amblyopia. Of particular interest may be the combination of rTMS and perceptual training since combining rTMS with other therapeutic interventions may be a promising therapeutic route (Ridding, M. C, and Rothwell, J. C. 2007. Nat Rev Neurosci. 8, 559-567).

Figures 8 and 9 are presented as complements to the data of Figures 1 and 2, respectively, but with additional subjects. In Figure 8, seven of nine patients had responded to the stimulation at one or both of the two post-rTMS time points, and if these results were considered alone, the effect of rTMS was reliable at T2 in Figure 8B. However, applicants have not been able to identify any distinguishing features for the non responders that would allow applicants to consider them as a clearly separate population. Interestingly, although the magnitude of the change was small, seven of nine and eight of nine

participants showed a reduction in contrast sensitivity at T1 and T2, respectively, for the non amblyopic eye, high spatial frequency condition presented in Figures 8A and 8B. The reduction was reliable for T2. No other conditions showed reliable rTMS-induced changes. Individual data are shown in Figures 8C and 8D for T1 and 12, respectively, plotted as the absolute change from the baseline in the amblyopic eye as a function of the difference in baseline performance between the two eyes (amblyopic eye pre-rTMS baseline 2 fellow fixing eye pre-rTMS baseline). The non responding participants are shown in gray. The positive correlations (marginal at T1 , reliable at T2) suggest that the larger the absolute difference between the two eyes at the baseline, the greater the effect of rTMS on the amblyopic eye. For 10 Hz stimulation, the result was clearer; all six participants tested showed improved contrast sensitivity at T1 and T2, as shown in Figure 9A. Importantly, both participants that did not respond to the 1 Hz stimulation did show a response to the 10 Hz stimulation. Figures 9B and 9C show that the absolute amount of improvement was positively correlated with the difference in the baseline between the two eyes. Although this correlation was driven predominantly by the most extreme data point, the pattern is consistent with the 1 Hz data.

A comparison of Figures 8C and 8D with Figures 9B and 9C shows a difference in the baselines between the two conditions for the most extreme data point (participant A.M.). This participant had to be tested at different spatial frequencies in the 10 Hz condition because of a sustained improvement in contrast sensitivity in the amblyopic eye after the 1 Hz rTMS. This improvement cannot be attributed only to the rTMS intervention, however, because A.M. had been recruited for a perceptual-training experiment in the intervening time between rTMS sessions. For all other participants, there was no significant change in baseline sensitivity across the different stimulation sessions (p > 0.05), which were separated by at least 1 week, indicating that the effects of rTMS were transient. Delivery of 1 Hz rTMS over motor cortex elicited no reliable changes in contrast sensitivity for the amblyopic observers (data not shown).

For all amblyopic participants, 10 Hz rTMS over visual cortex improved contrast detection for high spatial frequencies in the amblyopic eye directly after and 30 min after rTMS. The 1 Hz rTMS had less consistent effects, although the data suggest that this intervention may also be effective if the difference in function between the eyes is large. Inter-subject variability is a documented phenomenon in rTMS studies, particularly with shorter stimulation trains, and may therefore have been a factor here for the 1 Hz -stimulation paradigm. With applicant's currently evolving but incomplete understanding of rTMS, it is not possible to conclusively identify the mechanisms responsible for the rTMS-based improvement in visual function that reported here. However, applicants can assume that explanations based simply on global excitation or inhibition are unlikely to be satisfactory because both 1 Hz and 10 Hz stimulation were effective in the majority of subjects. This, therefore, implicates mechanisms requiring either (1 ) more complex changes in the relative excitation and inhibition of separate neural populations or (2) changes in ICI. The most parsimonious explanation is that rTMS acts to equate the excitability of the neurons subserving each eye. The direction of the change in the relative excitability of the populations of these neurons is still an open question. Although 1 Hz stimulation is thought to decrease excitability, it has been demonstrated that if a neural population is inhibited prior to rTMS (as may be the case for amblyopic-eye neurons), the effects of subsequent rTMS can be reversed.

Despite these considerations, the concept of promoting equality in neural excitability between the two eyes is still consistent with the efficacy of both 1 Hz and 10 Hz rTMS demonstrated in this study and the idea that rTMS preferentially acts to return a neural system to equilibrium (Ridding and Rothwell, 2007). This explanation is also partially supported by the small but reliable reduction in sensitivity in the fellow fixing eye after 1 Hz rTMS; i.e., rTMS had opposite effects on the two eyes of amblyopes for high spatial frequencies. A similar but not statistically reliable trend was also present in the 10 Hz data. A second possibility is that rTMS acts to reduce ICI, an effect that

has been demonstrated in the motor cortex for both 1 Hz and 10 Hz stimulation. In motor cortex, higher stimulation frequencies do appear to be more effective at modulating ICI. Such an effect may account for the finding that for two participants, 10 Hz stimulation was more effective than 1 Hz stimulation. An explanation based on ICI would link results with recent animal investigations highlighting the importance of reductions of ICI to recovery from visual deprivation (Sale et al. 2007). Unfortunately, it is not possible to measure ICI in visual cortex with a subjective phosphene report.

The nature of the cortical deficiency in amblyopia shares many features with stroke including the suppressive influence of one population of neurons over another, "weaker" population. In a healthy brain the two hemispheres achieve a state of balanced activity by mutual inhibition through the corpus callosum. Early after stroke, the lesioned hemisphere is no longer able to contribute to normal transcallosal inhibition and the unaffected hemisphere becomes hyperactive. An increase in the transcallosal inhibitory influence coming from the hyperactive hemisphere then inhibits the damaged hemisphere still further This inhibitory effect interferes with recovery of functions of the lesioned area in stroke.

Evidence for the important role of neuronal excitability in functional motor recovery provides a strong rationale for the development of therapeutic interventions aimed at manipulating the inter-hemispheric balance of excitability. Repetitive transcranial magnetic stimulation (rTMS) has recently been employed as a method to rectify the imbalance in inhibition between the two hemispheres either by driving down excitability in the dominant "inhibiting" hemisphere or driving up excitability in the "inhibited" lesioned hemisphere. The effects of rTMS are likely to be mediated by altered synaptic transmission in the cortex, as seen in the well-known phenomena of long-term potentiation and depression. There is evidence to suggest that the excitability of the stimulated region remains altered for a period of time after the offset of stimulation, with low stimulation frequencies (e.g. 1 Hz) decreasing excitability and higher frequencies (10-20 Hz) increasing excitability.

Extrapolating from the use of rTMS in stroke it is logical to employ the same principles to amblyopia where one population of dominant neurons is thought to be inhibiting a second population of "amblyopic" neurons. The delivery of rTMS is complicated however by the fact that, unlike the motor system, the visual system is lateralised in terms of visual space where the left side of the visual field is represented on the retinae of both eyes and processed by the right hemisphere and vice-versa. Therefore information from both eyes is processed within a single hemisphere. As previously mentioned, there is now a rising consensus from studies of both motor and visual cortex that the effects of TMS are critically dependent on the current activity state of the stimulated neurons in the cortex. The principle of homeostatic plasticity, first described in studies of synaptic plasticity, applies also to the effects of rTMS: the effects of excitatory stimulation are greatest in a system that has previously been inhibited, and vice versa (Ridding and Rothwell, 2007). This interaction between the current level of activity in a neural population and the effects of rTMS may allow applicants to influence the function from amblyopic eye and non-amblyopic eye neurons independently within a single stimulated region of visual cortex, since amblyopic and non-amblyopic eye neurons differ in their levels of excitability. The results from the application of rTMS to stroke recovery are promising (Wassermann, E. M. 1998. Electroencephalogr Clin Neurophysiol 108, 1-16). Here, applicants present data showing that repetitive transcranial magnetic stimulation (rTMS) of the visual cortex can temporarily improve contrast sensitivity in the amblyopic visual cortex. These results represent the first application of rTMS to the problem of amblyopia and demonstrate a direct transfer of insights into the abnormal inhibitory interactions gained in stroke, applied to amblyopia.

Methods

Psychophysics

Contrast sensitivity was measured using single, 17° Gabor patches presented within a 1 second temporal envelope. Patients had to indicate whether the

patches were oriented vertically or horizontally. Thresholds were measured using a 2 up 1 down staircase technique comprising six reversals (incorrect responses only), the last five of which were averaged to obtain the threshold. This procedure was repeated at least three times for each eye/spatial frequency combination in a random order within one 'block' of measurements, i.e. t1 , t2 and t3. The average of these measurements was used as the patient's threshold for that block. Stimuli were presented on a linearized lyama Vision Master Pro monitor using a ViSaGe visual stimulus generator (Cambridge Research Systems, UK). Patients performed the psychophysical task monocularly. An eye patch was used to occlude one eye.

Single pulse TMS

Phosphene thresholds (the stimulation intensity required to evoke a phosphene - a visual event such as a brief flash that is not induced by light hitting the retina) were acquired over V1 and subsequently used to calibrate the intensity used during the rTMS administration. During this procedure participants were asked to wear a white swimming cap so that certain points from the international ten-twenty electrode system could be marked on their head. In addition, during this step, participants were asked to wear blacked out swimming goggles so that light to their eyes was blocked, but they could still comfortably keep their eyes open. In order to localize V1 the coil was placed at electrode site O1 (occipital pole) and single pulse stimulation at 50- 100% stimulator output was applied over a 1x1 cm ² interval grid centered on O1. To verify reported phosphenes as being due to visual cortex stimulation the coil was moved laterally to check that the reported phosphene moved contra-laterally. In addition applicants ensured that no phosphenes were reported after stimulation of non-visual control site OZ located at the top of the head. Participants were asked to report the presence of static phosphenes and the site of stimulation that generated the strongest phosphene percept was chosen for the rTMS procedure. Once the optimal phosphene site was located, a phosphene threshold was measured, defined as the lowest amount of stimulation that gave rise to the percept of a phosphene on five out of ten

pulses. This procedure typically took approximately 30 minutes. To avoid effects of dark adaptation and to avoid discomfort from the light proof goggles patients wore during this procedure, patients were re-exposed to light at convenient intervals throughout testing. For motor cortex, a region of cortex in the right hemisphere corresponding to primary motor cortex was stimulated with single pulses until a twitch in the relaxed left FDI muscle was either reported by the patient or observed by the second experimenter. The location that produced the largest subjective twitch was located and then stimulation intensity was reduced until 5/10 pulses elicited the subjective report of a twitch. Subjective report was used as it paralleled the phosphene thresholding procedure used for visual cortex and therefore made the motor stimulation a more plausible control for the patients.

Repetitive TMS 1 Hz rTMS was delivered for 10 minutes (600 pulses) at 100% of threshold. 10 Hz rTMS was delivered to visual cortex only at 100% of motor threshold since this was consistently lower than phosphene threshold and therefore safer. 10 Hz rTMS was delivered in bursts of 5 second trains separated by 45 second inter train intervals (total 900 pulses). One patient did not undergo the motor control condition but did show differential responses to 1 Hz and 10Hz visual cortex stimulation supporting the absence of a non-specific rTMS related confound. Of the 6 patents that completed the motor control condition, 4 were stimulated over visual cortex on the first session. During the study one participant withdrew early in the first phosphene localization session as they did not tolerate the sensation of single pulse stimulation. No other participants reported any adverse effects of the stimulation. All procedures were approved by the institutional ethics committee. TMS was administered using a MagStim Rapid2 biphasic stimulator and a MagStim figure-8 air-cooled coil. During rTMS administration the BrainSight Frameless ^® stereotaxic system was used to monitor coil position to keep the position constant.

Differences between 1 Hz and 10 Hz Stimulation Parameters

Phosphene thresholds were measured using single pulse stimulation resulting in relatively high thresholds (mean 85% maximum stimulator output, SD 7% for amblyopes and mean 72% Maximal Stimulation Output (MSO), SD 10% for controls). Initial pilot observations identified tolerability issues for 1 Hz stimulation over 600 pulses, so we adopted 600 pulses as our train length, a duration that has previously been shown to be effective. 10Hz stimulation was intolerable at 100% single pulse phosphene threshold (lower absolute intensities have been used elsewhere), we therefore used 100% motor threshold since these were lower than phosphene thresholds (mean 69% MSO, SD 12% for amblyopes, mean 59% for controls, SD 10%). To mitigate against the reduction in intensity, we adopted a longer pulse train (900 pulses). Interestingly, we observed a significant positive correlation across participants between single pulse phosphene thresholds and subjective motor thresholds (rho = 0.56, p < 0.05) with motor threshold representing 82% of phosphene threshold on average (SD = 13%).