Microneurography and Intraneural Microstimulation for Upper Limb Sensory Investigation in Healthy and Amputee Subjects

Experimental Study to Investigate Sensory Control of the Upper Limb Through Microneurography and to Restore Near-natural Tactile Sensations Through Intraneural Microstimulation in Healthy and Amputee Subjects

The goal of this experimental study is to investigate the somatosensory system through intraneural recording and stimulation of the median nerve in healthy volunteers and participants with upper-limb amputation. The main questions it aims to answer are:

• How does median nerve activity encode tactile perception during different types of mechanical tactile stimulation in healthy subjects?
• How can tactile information be artificially encoded and restored through intraneural microstimulation in both healthy subjects and upper-limb amputee participants? How rich and functionally meaningful is the restored sensory information?
• What are the neurophysiological mechanisms underlying tactile stimulus processing and the associated patterns of brain activation during tactile stimulation?

Participants will undergo median nerve recordings using microneurography during controlled mechanical tactile stimulation, as well as intraneural microstimulation with specific patterns to evoke near-natural tactile sensations. They will perform perceptual reporting tasks related to stimulus type, intensity, and localization, as well as psychophysical tests under different stimulation conditions. When possible, they will also undergo non-invasive recordings of brain activity using electroencephalography during sensory stimulation.

Participants will attend the laboratory where the entire research protocol will be conducted in a single visit lasting up to 4 hours.

Medical history and a physical examination will be collected. In participants with amputation, additional clinical information from previous neurological assessments will be reviewed (including medical records, nerve conduction velocity data, and ultrasound or MRI examinations required to verify inclusion/exclusion criteria). Psychometric tests will also be administered to assess language abilities, visuospatial skills, and general intelligence, ensuring that participants are able to understand and perform the experimental tasks. If inclusion and exclusion criteria are met, participants will undergo microneurography and/or intraneural microstimulation procedures. Participants will be seated comfortably, with the dominant arm (healthy volunteers) or the residual limb (participants with amputation) supported. A trained medical operator will identify the median nerve using external stimulation and/or ultrasound guidance to assist electrode insertion. The active electrode will be inserted into the nerve, while a reference electrode will be placed in the subcutaneous tissue a few centimeters away. Correct electrode placement will then be refined using intraneural stimulation. Once the nerve fascicle is reached, tactile stimulation of the hand will be used to identify single mechanoreceptive units. Final electrode placement will be confirmed by visual and auditory inspection of the neural signal. The median nerve activity must be clearly distinguishable from background activity when mechanical stimuli are applied to the skin. During electrode insertion, participants will be instructed to remain still until the procedure is completed. The intraneural microstimulation session, performed in both healthy volunteers and participants with upper-limb amputation, is preceded by peripheral neural recordings using microneurography during the presentation of tactile stimuli (e.g., Von Frey filaments). This step is used for accurate nerve localization and preliminary assessment of the receptors and their receptive fields. Once the neural target is identified, participants undergo intraneural stimulation involving the perception and recognition of artificially generated tactile stimuli. These stimuli are delivered via intraneural microstimulation and are derived from mechanoneural transduction, using a sensorized artificial finger interacting with different surfaces to encode surface-dependent tactile information. Participants will be asked to recognize different properties of the presented stimuli. During the tests, electroencephalographic (EEG) activity may be recorded using a 128-channel EEG system, synchronized with the onset of stimulation.

Inclusion Criteria:

Healthy participants

• Age between 18-60 years, of any sex
• Signed informed consent
• No neurological diseases
• Presence of both intact upper limbs
• Normal language, visuospatial, and general intellectual abilities, assessed using the Wechsler Adult Intelligence Scale-Revised (WAIS-R) (Appendix 1)

Amputee participants

• Age between 18-60 years, of any sex
• Signed informed consent
• Upper-limb amputation (transradial or hand amputation)
• Stable amputation for at least one year
• Normal language, visuospatial, and general intellectual abilities, assessed using the Wechsler Adult Intelligence Scale-Revised (WAIS-R) (Appendix 1)
• Evidence of residual peripheral nerve function in the stump, based on nerve conduction studies as documented in clinical records

Exclusion Criteria:

Healthy participants

• Conditions affecting perceptual abilities
• Inability to provide informed consent
• Pregnancy or breastfeeding (self-reported)
• Clinical conditions that, in the judgment of the investigator responsible for recruitment, may interfere with microneurography, microstimulation, and EEG procedures
• Current or past substance abuse
• Brain injury with residual motor impairment
• Depression
• Presence of neurological or musculoskeletal disorders
• Diabetes mellitus
• Current or previous dermatological conditions
• Morphological nerve abnormalities, as indicated by ultrasound or MRI findings in clinical records
• Upper-limb amputation

Amputee participants

• Psychological and/or cognitive disorders
• Inability to provide informed consent
• Pregnancy or breastfeeding (self-reported)
• Clinical conditions that, in the judgment of the investigator responsible for recruitment, may interfere with microneurography, microstimulation, and EEG procedures
• Current or past substance abuse
• Brain injury with residual motor impairment
• Depression
• Presence of neurological or musculoskeletal disorders
• Diabetes mellitus
• Current or previous dermatological conditions
• Morphological nerve abnormalities, as indicated by ultrasound or MRI findings in clinical records
• Bilateral upper-limb amputation