After establishing a 3-minute rolling baseline average, the hindbrain stimulator (upper right of Fig. 3A) was armed.
Electrical stimulation of the right supraorbital nerve was performed with a peripheral nerve stimulator and silver/silver chloride disc surface electrodes (DS7A Stimulator, Digitimer Ltd., Welwyn Garden City, Hertz, UK).
Ventricular fibrillation (VF) was induced by an electrical stimulator connected to the right ventricular pacing lead.
The vagus nerve stimulator can be activated right after surgery.
The latter two types of stimulation excited the right ventricle using constant voltage pulses (1 4 V, 2 ms duration; Grass S48 stimulator, Grass Technologies, West Warwick, RI, USA).
Individual TMS resting motor thresholds were obtained in a separate session, via stimulation over the right M1 'hotspot' for inducing a visible twitch of the first dorsal interosseus (mean resting motor threshold with conventional TMS coil 56 ± 10% of maximal stimulator output; or with the non-ferrous MR-compatible coil that had a longer cable 67±10%0% of maximal output).
The second point is that even when using the right amount of stimulator, the total number of stimulations is also important.
Since it gradually became impossible to achieve sufficient ventilation by left phrenic nerve stimulation, probably due to stimulator detachment, the output strength of the IPG for right phrenic nerve stimulation and the duration of stimulation were managed to achieve a target EtCO2 level of 50 60 mmHg.
After the tactile stimulators were connected to their right index finger and the ear plugs were inserted, a calibration was performed to find the participants approximate detection threshold of the experimental stimuli.
An allodynia score was determined after four consecutive measurements using the touch stimulator sequentially on the left and right hind paw and calculated considering the formula below: The basal score was measured before CFA injection on day 0, and the animals considered for testing were those with a mean value nearest to 1 (demonstrating no significant difference between both paw stimuli).
