This model can become a standard for safety and efficacy validation for chronic cortical devices in terms of biocompatibility, resolution and signal quality. Our procedure describes a reproducible and scalable method to perform long-term follow-up of device safety and efficacy. This includes neural monitoring over time, as well as in vivo and ex vivo imaging.
Our method will help in the development of sensory and motor cortical neuroprosthesis. It is a tool to understand the large scale cortical networks activity. This method can be used in fundamental neuroscience to investigate functional connectivity over different cortical areas.
Can also be applied to other large animal models. The surgical approach needs some practice, which can be acquired on cadavers or acute experiments first. The measurements should then be fairly straightforward.
To begin, incise the skin of the anesthetized animal with a scalpel knife along the midline. Separate the muscle and periosteum from the bone using a raspatory and place spreaders for optimum access. To perform the craniotomy, drill the outline using a bone drill with a round cutting bit, taking into account the thickness of the skull as measured on the X-ray.
Irrigate the drilling location with saline solution to avoid overheating the bone. Carefully drill the outline homogeneously until reaching the dura mater. At the first breakthrough, finish drilling the outline until it has thinned enough to almost break through.
Then use a flat spatula to break away the bone flap in one piece using the craniotomy edge as leverage. To perform durotomy, using the needle from a six oh suture kit, carefully pierce and lift the dura mater at the anterior or posterior end of the craniotomy, halfway between the medial and lateral side, and create the beginning of an incision with the stab knife. Then using a small flat spatula inserted in the subdural space, acting as a cutting base to protect the cortex.
Create an anteroposterior slit in the dura mater by advancing simultaneously with both tools. Ensure that the slit is slightly larger than the width of the implant. Place the implant above the dura mater slit and with small forceps, subdurally insert the device by sliding it sequentially on each edge.
Carefully hold the pedestal end of the device and advance with the implant in order not to create tension hindering the insertion. Stop the insertion when the connector edge is located on top of the slit. To secure the implant in place, place a titanium bridge over the cable after the edge of the craniotomy or in the anchoring wings, and secure it with one or two titanium screws using the appropriate screwdriver.
Next, suture the dura mater carefully around the implant cable. Using a three oh resorbable suture and a small needle holder, bring the two dura mater edges together as much as possible without tearing through the thin membrane with the suture wire. To perform bone flap placement, fix a titanium bridge on the anterior and posterior part of each bone flap using a titanium screw.
Screw the end of the titanium bridges to the skull. Next, plan the orientation of the footplate to ensure all legs can be screwed into the skull. Then secure the footplate by screwing in the titanium screws of the footplate until it is firmly in place.
Then screw the pedestal onto the footplate. Create subcutaneous sutures with a four oh non-resorbable suture wire with sutures three millimeters apart. Start away from the pedestal, moving toward it on both sides of the incision.
Next, close the dermal layer by suturing the skin using a six oh non-resorbable suture wire. With sutures five millimeters apart. Start away from the pedestal, moving toward it on both sides of the incision.
Take care to achieve good tissue apposition between the two skin flaps and near the pedestal edge to avoid a void. After plugging the wireless head stage onto the animal achieved by either holding the animal or distracting it by feeding with treats, record the awake brain signals. Ensure to place the amplifier antenna and the external speakers close to the pig's cage while recording the signals.
Baseline activity without sound stimuli and auditory evoked potentials in response to an 800 hertz tone burst stimulation can be mapped over the electrode array. The auditory evoked potential in a single electrode channel is shown over time with arrows marking the on"response and the baseline activity is shown as a comparison. In vivo imaging was performed intraoperatively and postoperatively to assess the brain state and implant positioning.
Intraoperative plane x-ray verified implant placement, and no folding as observed by the radiopaque marker placement. The brain's surface is intact as can be observed in the postoperative MRI. Overall, with this implant and pedestal system, whole brain imaging is possible over the course of the implantation period to see anatomical structures or the presence of liquid and blood around the implant.
Additionally, clinical electrodes are used as comparators in this study, but cannot be imaged in the MRI due to heating and safety concerns and require CT scans. The presented pipeline enables whole brain extraction and sectioning to image whole hemispheres. The imaging of the whole tissue section showed a clear neuron layer.
The cells are clearly defined on confocal imaging at 20x and enable fine investigation of inflammatory markers. Electrochemical characterization of the devices was used to extract the impedance modulus and phase in vitro, which was tracked over time at one kilohertz during the six months of implantation. It is crucial to carefully select the age and the size of the animal to avoid opening sinuses during the surgery.
This would compromise the chronic experiment. It is important to avoid bleeding when accessing the dura mater or inserting the implants. This will avoid further complications and inflammatory response.
Once this model is in place, it can be used to perform freely behaving electrophysiology in mini pigs and record activity from cortical areas of interest. This method can be applied in gathering biosafety data for submission of a clinical trial when developing new neuroprosthetics that will be translated to humans.
