The spinal cord facilitates cerebellar upper limb motor learning and control; inputs from neuromusculoskeletal simulation.

Complex interactions between brain regions and the spinal cord (SC) govern body motion, which is ultimately driven by muscle activation. Motor planning or learning are mainly conducted at higher brain regions, whilst the SC acts as a brain-muscle gateway and as a motor control centre providing fast reflexes and muscle activity regulation. Thus, higher brain areas need to cope with the SC as an inherent and evolutionary older part of the body dynamics. Here, we address the question of how SC dynamics affects motor learning within the cerebellum; in particular, does the SC facilitate cerebellar motor learning or constitute a biological constraint? We provide an exploratory framework by integrating biologically plausible cerebellar and SC computational models in a musculoskeletal upper limb control loop. The cerebellar model, equipped with the main form of cerebellar plasticity, provides motor adaptation; whilst the SC model implements stretch reflex and reciprocal inhibition between antagonist muscles. The resulting spino-cerebellar model is tested performing a set of upper limb motor tasks, including external perturbation studies. A cerebellar model, lacking the implemented SC model and directly controlling the simulated muscles, was also tested in the same. The performances of the spino-cerebellar and cerebellar models were then compared, thus allowing directly addressing the SC influence on cerebellar motor adaptation and learning, and on handling external motor perturbations. Performance was assessed in both joint and muscle space, and compared with kinematic and EMG recordings from healthy participants. The differences in cerebellar synaptic adaptation between both models were also studied. We conclude that the SC facilitates cerebellar motor learning; when the SC circuits are in the loop, faster convergence in motor learning is achieved with simpler cerebellar synaptic weight distributions. The SC is also found to improve robustness against external perturbations, by better reproducing and modulating muscle cocontraction patterns.

Walking naturally after spinal cord injury using a brain-spine interface.

A spinal cord injury interrupts the communication between the brain and the region of the spinal cord that produces walking, leading to paralysis1,2. Here, we restored this communication with a digital bridge between the brain and spinal cord that enabled an individual with chronic tetraplegia to stand and walk naturally in community settings. This brain-spine interface (BSI) consists of fully implanted recording and stimulation systems that establish a direct link between cortical signals3 and the analogue modulation of epidural electrical stimulation targeting the spinal cord regions involved in the production of walking4-6. A highly reliable BSI is calibrated within a few minutes. This reliability has remained stable over one year, including during independent use at home. The participant reports that the BSI enables natural control over the movements of his legs to stand, walk, climb stairs and even traverse complex terrains. Moreover, neurorehabilitation supported by the BSI improved neurological recovery. The participant regained the ability to walk with crutches overground even when the BSI was switched off. This digital bridge establishes a framework to restore natural control of movement after paralysis.

Activity-dependent spinal cord neuromodulation rapidly restores trunk and leg motor functions after complete paralysis.

Epidural electrical stimulation (EES) targeting the dorsal roots of lumbosacral segments restores walking in people with spinal cord injury (SCI). However, EES is delivered with multielectrode paddle leads that were originally designed to target the dorsal column of the spinal cord. Here, we hypothesized that an arrangement of electrodes targeting the ensemble of dorsal roots involved in leg and trunk movements would result in superior efficacy, restoring more diverse motor activities after the most severe SCI. To test this hypothesis, we established a computational framework that informed the optimal arrangement of electrodes on a new paddle lead and guided its neurosurgical positioning. We also developed software supporting the rapid configuration of activity-specific stimulation programs that reproduced the natural activation of motor neurons underlying each activity. We tested these neurotechnologies in three individuals with complete sensorimotor paralysis as part of an ongoing clinical trial ( www.clinicaltrials.gov identifier NCT02936453). Within a single day, activity-specific stimulation programs enabled these three individuals to stand, walk, cycle, swim and control trunk movements. Neurorehabilitation mediated sufficient improvement to restore these activities in community settings, opening a realistic path to support everyday mobility with EES in people with SCI.

Recruitment of upper-limb motoneurons with epidural electrical stimulation of the cervical spinal cord.

Epidural electrical stimulation (EES) of lumbosacral sensorimotor circuits improves leg motor control in animals and humans with spinal cord injury (SCI). Upper-limb motor control involves similar circuits, located in the cervical spinal cord, suggesting that EES could also improve arm and hand movements after quadriplegia. However, the ability of cervical EES to selectively modulate specific upper-limb motor nuclei remains unclear. Here, we combined a computational model of the cervical spinal cord with experiments in macaque monkeys to explore the mechanisms of upper-limb motoneuron recruitment with EES and characterize the selectivity of cervical interfaces. We show that lateral electrodes produce a segmental recruitment of arm motoneurons mediated by the direct activation of sensory afferents, and that muscle responses to EES are modulated during movement. Intraoperative recordings suggested similar properties in humans at rest. These modelling and experimental results can be applied for the development of neurotechnologies designed for the improvement of arm and hand control in humans with quadriplegia.

Local photoreceptor degeneration causes local pathophysiological remodeling of retinal neurons.

Vision loss in age-related macular degeneration (AMD) stems from disruption of photoreceptor cells in the macula, the central retinal area required for high-acuity vision. Mice and rats have no macula, but surgical insertion of a subretinal implant can induce localized photoreceptor degeneration due to chronic separation from retinal pigment epithelium, simulating a key aspect of AMD. We find that the implant-induced loss of photoreceptors in rat retina leads to local changes in the physiology of downstream retinal ganglion cells (RGCs), similar to changes in RGCs of rodent models of retinitis pigmentosa (RP), an inherited disease causing retina-wide photoreceptor degeneration. The local implant-induced changes in RGCs include enhanced intrinsic excitability leading to accelerated spontaneous firing, increased membrane permeability to fluorescent dyes, and enhanced photosensitization by azobenzene photoswitches. The local physiological changes are correlated with an increase in retinoic acid receptor-induced (RAR-induced) gene transcription, the key process underlying retinal remodeling in mouse models of RP. Hence the loss of photoreceptors, whether by local physical perturbation or by inherited mutation, leads to a stereotypical set of pathophysiological consequences in RGCs. These findings implicate RAR as a possible common therapeutic target for reversing the signal-corrupting effects of retinal remodeling in both RP and AMD.

Behavioural responses to a photovoltaic subretinal prosthesis implanted in non-human primates.

Retinal dystrophies and age-related macular degeneration related to photoreceptor degeneration can cause blindness. In blind patients, although the electrical activation of the residual retinal circuit can provide useful artificial visual perception, the resolutions of current retinal prostheses have been limited either by large electrodes or small numbers of pixels. Here we report the evaluation, in three awake non-human primates, of a previously reported near-infrared-light-sensitive photovoltaic subretinal prosthesis. We show that multipixel stimulation of the prosthesis within radiation safety limits enabled eye tracking in the animals, that they responded to stimulations directed at the implant with repeated saccades and that the implant-induced responses were present two years after device implantation. Our findings pave the way for the clinical evaluation of the prosthesis in patients affected by dry atrophic age-related macular degeneration.

Retinal Laser Therapy Preserves Photoreceptors in a Rodent Model of MERTK-Related Retinitis Pigmentosa.

PURPOSE: We investigated the effects of various retinal laser therapies on preservation of the photoreceptors in an animal model of Mer tyrosine kinase receptor (MERTK)-related retinitis pigmentosa (RP). These modalities included photocoagulation with various pattern densities, selective RPE therapy (SRT), and nondamaging retinal therapy (NRT). METHODS: Laser treatments were performed on right eyes of RCS rats, using one of three laser modalities. For photocoagulation, six pattern densities (spot spacings of 0.5, 1, 1.5, 3, 4, and 5 spot diameters) were delivered in 19-day-old animals, prior to the onset of photoreceptor degeneration, to determine the optimal treatment density for the best preservation of photoreceptors. The left eye was used as control. Rats were monitored for 6 months after treatment using electroretinography, optical coherence tomography, and histology. RESULTS: Photocoagulation resulted in long-term preservation of photoreceptors, manifested morphologically and functionally, with the extent of the benefit dependent on the laser pattern density. Eyes treated with a 1.5 spot size spacing showed the best morphologic and functional preservation during the 6-month follow-up. SRT-treated eyes exhibited short-term morphologic preservation, but no functional benefit. NRT-treated eyes did not show any observable preservation benefit from the treatment. CONCLUSIONS: In a rodent model of MERTK-related RP, pattern photocoagulation of about 15% of the photoreceptors (1.5 spot diameter spacing) provides long-term preservation of photoreceptors in the treatment area. TRANSLATIONAL RELEVANCE: Availability of retinal lasers in ophthalmic practice enables rapid translation of our study to clinical testing and may help preserve the sight in patients with photoreceptor degeneration.

Characteristics of prosthetic vision in rats with subretinal flat and pillar electrode arrays.

OBJECTIVE: Retinal prostheses aim to restore sight by electrically stimulating the surviving retinal neurons. In clinical trials of the current retinal implants, prosthetic visual acuity does not exceed 20/550. However, to provide meaningful restoration of central vision in patients blinded by age-related macular degeneration (AMD), prosthetic acuity should be at least 20/200, necessitating a pixel pitch of about 50 µm or lower. With such small pixels, stimulation thresholds are high due to limited penetration of electric field into tissue. Here, we address this challenge with our latest photovoltaic arrays and evaluate their performance in vivo. APPROACH: We fabricated photovoltaic arrays with 55 and 40 µm pixels (a) in flat geometry, and (b) with active electrodes on 10 µm tall pillars. The arrays were implanted subretinally into rats with degenerate retina. Stimulation thresholds and grating acuity were evaluated using measurements of the visually evoked potentials (VEP). MAIN RESULTS: With 55 µm pixels, we measured grating acuity of 48 ± 11 µm, which matches the linear pixel pitch of the hexagonal array. This geometrically corresponds to a visual acuity of 20/192 in a human eye, matching the threshold of legal blindness in the US (20/200). With pillar electrodes, the irradiance threshold was nearly halved, and duration threshold reduced by more than three-fold, compared to flat pixels. With 40 µm pixels, VEP was too low for reliable measurements of the grating acuity, even with pillar electrodes. SIGNIFICANCE: While being helpful for treating a complete loss of sight, current prosthetic technologies are insufficient for addressing the leading cause of untreatable visual impairment-AMD. Subretinal photovoltaic arrays may provide sufficient visual acuity for restoration of central vision in patients blinded by AMD.

Transplantation of Mature Photoreceptors in Rodents With Retinal Degeneration.

PURPOSE: To demonstrate survival and integration of mature photoreceptors transplanted with the retinal pigment epithelium (RPE). METHODS: Full-thickness retina with attached RPE was harvested from healthy adult rats. Grafts were implanted into two rat models of retinal degeneration, Royal College of Surgeons (RCS) and S334ter-3. Survival of the host and transplanted retina was monitored using optical coherence tomography (OCT) for up to 6 months. The retinal structure and synaptogenesis between the host and transplant was assessed by histology and immunohistochemistry. RESULTS: OCT and histology demonstrated a well-preserved photoreceptor layer with inner and outer segments, while the inner retinal layers of the transplant largely disappeared. Grafts, including RPE, survived better than without and the transplanted RPE appeared as a monolayer integrated with the native one. Synaptogenesis was observed through sprouting of new dendrites from the host bipolar cells and synaptic connections forming with cells of the transplant. However, in many samples, a glial fibrillary acidic protein-positive membrane separated the host retina and the graft. CONCLUSIONS: Presence of RPE in the graft improved the survival of transplanted photoreceptors. Functional integration between the transplant and the host retina is likely to be further enhanced if formation of a glial seal could be prevented. Transplantation of the mature photoreceptors with RPE may be a practical approach to restoration of sight in retinal degeneration. TRANSLATIONAL RELEVANCE: This approach to restoration of sight in patients with photoreceptor degeneration can be rapidly advanced to clinical testing. In patients with central scotoma, autologous transplantation of the peripheral retina can be an option.

Temporal structure in spiking patterns of ganglion cells defines perceptual thresholds in rodents with subretinal prosthesis.

Subretinal prostheses are designed to restore sight in patients blinded by retinal degeneration using electrical stimulation of the inner retinal neurons. To relate retinal output to perception, we studied behavioral thresholds in blind rats with photovoltaic subretinal prostheses stimulated by full-field pulsed illumination at 20 Hz, and measured retinal ganglion cell (RGC) responses to similar stimuli ex-vivo. Behaviorally, rats exhibited startling response to changes in brightness, with an average contrast threshold of 12%, which could not be explained by changes in the average RGC spiking rate. However, RGCs exhibited millisecond-scale variations in spike timing, even when the average rate did not change significantly. At 12% temporal contrast, changes in firing patterns of prosthetic response were as significant as with 2.3% contrast steps in visible light stimulation of healthy retinas. This suggests that millisecond-scale changes in spiking patterns define perceptual thresholds of prosthetic vision. Response to the last pulse in the stimulation burst lasted longer than the steady-state response during the burst. This may be interpreted as an excitatory OFF response to prosthetic stimulation, and can explain behavioral response to decrease in illumination. Contrast enhancement of images prior to delivery to subretinal prosthesis can partially compensate for reduced contrast sensitivity of prosthetic vision.

Optimization of pillar electrodes in subretinal prosthesis for enhanced proximity to target neurons.

OBJECTIVE: High-resolution prosthetic vision requires dense stimulating arrays with small electrodes. However, such miniaturization reduces electrode capacitance and penetration of electric field into tissue. We evaluate potential solutions to these problems with subretinal implants based on utilization of pillar electrodes. APPROACH: To study integration of three-dimensional (3D) implants with retinal tissue, we fabricated arrays with varying pillar diameter, pitch, and height, and implanted beneath the degenerate retina in rats (Royal College of Surgeons, RCS). Tissue integration was evaluated six weeks post-op using histology and whole-mount confocal fluorescence imaging. The electric field generated by various electrode configurations was calculated in COMSOL, and stimulation thresholds assessed using a model of network-mediated retinal response. MAIN RESULTS: Retinal tissue migrated into the space between pillars with no visible gliosis in 90% of implanted arrays. Pillars with 10 µm height reached the middle of the inner nuclear layer (INL), while 22 µm pillars reached the upper portion of the INL. Electroplated pillars with dome-shaped caps increase the active electrode surface area. Selective deposition of sputtered iridium oxide onto the cap ensures localization of the current injection to the pillar top, obviating the need to insulate the pillar sidewall. According to computational model, pillars having a cathodic return electrode above the INL and active anodic ring electrode at the surface of the implant would enable six times lower stimulation threshold, compared to planar arrays with circumferential return, but suffer from greater cross-talk between the neighboring pixels. SIGNIFICANCE: 3D electrodes in subretinal prostheses help reduce electrode-tissue separation and decrease stimulation thresholds to enable smaller pixels, and thereby improve visual acuity of prosthetic vision.

Col4a1 mutation generates vascular abnormalities correlated with neuronal damage in a mouse model of HANAC syndrome.

The HANAC syndrome is caused by mutations in the gene coding for collagen4a1, a major component of blood vessel basement membranes. Ocular symptoms include an increase in blood vessel tortuosity and occasional hemorrhages. To examine how vascular defects can affect neuronal function, we analyzed the retinal phenotype of a HANAC mouse model. Heterozygous mutant mice displayed both a thinning of the basement membrane in retinal blood vessels and in Bruch's membrane resulting in vascular leakage. Homozygous mice had additional vascular changes, including greater vessel coverage and tortuosity. This greater tortuosity was associated to higher expression levels of vascular endothelial growth factor (VEGF). These major changes to the blood vessels were correlated with photoreceptor dysfunction and degeneration. The neuronal damage was associated with reactive gliosis in astrocytes and Müller glial cells, and by the migration of microglial cells into the outer retina. This study illustrates how vascular changes can trigger neuronal degeneration in a new model of HANAC syndrome that can be used to further study dysfunctions of neurovascular coupling. SUMMARY STATEMENT: This study provides a phenotypic analysis of a novel mouse model of HANAC syndrome focusing on the retinal aspect. It recapitulates most of the aspects of the human disease and is therefore a great tool to study and to address this condition.

SiC protective coating for photovoltaic retinal prosthesis.

OBJECTIVE: To evaluate plasma-enhanced, chemically vapor deposited (PECVD) amorphous silicon carbide (α-SiC:H) as a protective coating for retinal prostheses and other implantable devices, and to study their failure mechanisms in vivo. APPROACH: Retinal prostheses were implanted in rats sub-retinally for up to 1 year. Degradation of implants was characterized by optical and scanning electron microscopy. Dissolution rates of SiC, SiN x and thermal SiO2 were measured in accelerated soaking tests in saline at 87 °C. Defects in SiC films were revealed and analyzed by selectively removing the materials underneath those defects. MAIN RESULTS: At 87 °C SiN x dissolved at 18.3 ± 0.3 nm d(-1), while SiO2 grown at high temperature (1000 °C) dissolved at 0.104 ± 0.008 nm d(-1). SiC films demonstrated the best stability, with no quantifiable change after 112 d. Defects in thin SiC films appeared primarily over complicated topography and rough surfaces. SIGNIFICANCE: SiC coatings demonstrating no erosion in accelerated aging test for 112 d at 87 °C, equivalent to about 10 years in vivo, can offer effective protection of the implants. Photovoltaic retinal prostheses with PECVD SiC coatings exhibited effective protection from erosion during the 4 month follow-up in vivo. The optimal thickness of SiC layers is about 560 nm, as defined by anti-reflective properties and by sufficient coverage to eliminate defects.

Implantation of Modular Photovoltaic Subretinal Prosthesis.

BACKGROUND AND OBJECTIVE: Establish the surgical procedure for subretinal implantation of multiple photovoltaic arrays for the restoration of sight. MATERIALS AND METHODS: Multiple silicon photovoltaic arrays of 1 mm in diameter and 30 µm in thickness were implanted subretinally via single retinotomy in rabbits. Ophthalmoscopic imaging and optical coherence tomography (OCT) were used to validate the implants' placement. RESULTS: Vitrectomy, followed by subretinal fluid injection for retinal detachment and retinotomy, allowed accurate placement of seven modules in the bleb, covering approximately a 3.5-mm diameter area on the retina via a single 1.5-mm retinotomy. OCT confirmed complete reattachment of the retina over the implants. CONCLUSION: Subretinal implantation of multiple photovoltaic arrays via a single retinotomy, followed by their tiling, minimizes the scleral and retinal incisions and provides better fit to the spherical shape of the eye ball, compared to a single, larger module. Such minimally traumatic procedure can be performed with 20-gauge intraocular instruments.

Interactions of Prosthetic and Natural Vision in Animals With Local Retinal Degeneration.

PURPOSE: Prosthetic restoration of partial sensory loss leads to interactions between artificial and natural inputs. Ideally, the rehabilitation should allow perceptual fusion of the two modalities. Here we studied the interactions between normal and prosthetic vision in a rodent model of local retinal degeneration. METHODS: Implantation of a photovoltaic array in the subretinal space of normally sighted rats induced local degeneration of the photoreceptors above the chip, and the inner retinal neurons in this area were electrically stimulated by the photovoltaic implant powered by near-infrared (NIR) light. We studied prosthetic and natural visually evoked potentials (VEP) in response to simultaneous stimulation by NIR and visible light patterns. RESULTS: We demonstrate that electrical and natural VEPs summed linearly in the visual cortex, and both responses decreased under brighter ambient light. Responses to visible light flashes increased over 3 orders of magnitude of contrast (flash/background), while for electrical stimulation the contrast range was limited to 1 order of magnitude. The maximum amplitude of the prosthetic VEP was three times lower than the maximum response to a visible flash over the same area on the retina. CONCLUSIONS: Ambient light affects prosthetic responses, albeit much less than responses to visible stimuli. Prosthetic representation of contrast in the visual scene can be encoded, to a limited extent, by the appropriately calibrated stimulus intensity, which also depends on the ambient light conditions. Such calibration will be important for patients combining central prosthetic vision with natural peripheral sight, such as in age-related macular degeneration.

Development of Animal Models of Local Retinal Degeneration.

PURPOSE: Development of nongenetic animal models of local retinal degeneration is essential for studies of retinal pathologies, such as chronic retinal detachment or age-related macular degeneration. We present two different methods to induce a highly localized retinal degeneration with precise onset time, that can be applied to a broad range of species in laboratory use. METHODS: A 30-µm thin polymer sheet was implanted subretinally in wild-type (WT) rats. The effects of chronic retinal separation from the RPE were studied using histology and immunohistochemistry. Another approach is applicable to species with avascular retina, such as rabbits, where the photoreceptors and RPE were thermally ablated over large areas, using a high power scanning laser. RESULTS: Photoreceptors above the subretinal implant in rats degenerated over time, with 80% of the outer nuclear layer disappearing within a month, and the rest by 3 months. Similar loss was obtained by selective photocoagulation with a scanning laser. Cells in the inner nuclear layer and ganglion cell layer were preserved in both cases. However, there were signs of rewiring and decrease in the size of the bipolar cell terminals in the damaged areas. CONCLUSIONS: Both methods induce highly reproducible degeneration of photoreceptors over a defined area, with complete preservation of the inner retinal neurons during the 3-month follow-up. They provide a reliable platform for studies of local retinal degeneration and development of therapeutic strategies in a wide variety of species.

Photovoltaic restoration of sight with high visual acuity.

Patients with retinal degeneration lose sight due to the gradual demise of photoreceptors. Electrical stimulation of surviving retinal neurons provides an alternative route for the delivery of visual information. We demonstrate that subretinal implants with 70-µm-wide photovoltaic pixels provide highly localized stimulation of retinal neurons in rats. The electrical receptive fields recorded in retinal ganglion cells were similar in size to the natural visual receptive fields. Similarly to normal vision, the retinal response to prosthetic stimulation exhibited flicker fusion at high frequencies, adaptation to static images and nonlinear spatial summation. In rats with retinal degeneration, these photovoltaic arrays elicited retinal responses with a spatial resolution of 64 ± 11 µm, corresponding to half of the normal visual acuity in healthy rats. The ease of implantation of these wireless and modular arrays, combined with their high resolution, opens the door to the functional restoration of sight in patients blinded by retinal degeneration.

Performance of photovoltaic arrays in-vivo and characteristics of prosthetic vision in animals with retinal degeneration.

Loss of photoreceptors during retinal degeneration leads to blindness, but information can be reintroduced into the visual system using electrical stimulation of the remaining retinal neurons. Subretinal photovoltaic arrays convert pulsed illumination into pulsed electric current to stimulate the inner retinal neurons. Since required irradiance exceeds the natural luminance levels, an invisible near-infrared (915 nm) light is used to avoid photophobic effects. We characterized the thresholds and dynamic range of cortical responses to prosthetic stimulation with arrays of various pixel sizes and with different number of photodiodes. Stimulation thresholds for devices with 140 µm pixels were approximately half those of 70 µm pixels, and with both pixel sizes, thresholds were lower with 2 diodes than with 3 diodes per pixel. In all cases these thresholds were more than two orders of magnitude below the ocular safety limit. At high stimulation frequencies (>20 Hz), the cortical response exhibited flicker fusion. Over one order of magnitude of dynamic range could be achieved by varying either pulse duration or irradiance. However, contrast sensitivity was very limited. Cortical responses could be detected even with only a few illuminated pixels. Finally, we demonstrate that recording of the corneal electric potential in response to patterned illumination of the subretinal arrays allows monitoring the current produced by each pixel, and thereby assessing the changes in the implant performance over time.