Diminished signal-to-noise ratio disrupts somatosensory population encoding and drives tactile hyposensitivity in the Fmr1-/y autism model.

Experimental design We reverse-translated a task used in human studies (Puts et al., 2014) and developed a novel Go/No-go detection task for vibrotactile stimuli which we combined with two-photon calcium imaging of excitatory and inhibitory neurons in the forepaw-related primary somatosensory cortex (FP-S1), to study altered tactile perception in autism and define its neural correlates.MiceSecond-generation Fmr1 knockout (Fmr1−/y) (Mientjes et al., 2006) and wild-type littermate mice, 5-16 weeks old, were used. Mice were fully congenic on a C57Bl/6J background (backcrossed for >10 generations into C57Bl/6J). Male wild-type (Fmr1+/y) and knockout (Fmr1−/y) littermates were generated by crossing Fmr1+/− females with Fmr1+/y male mice from the same colony. Post-weaning, mice were group-housed (2–4 males per cage), balanced for genotype, and provided cotton nestlets and carton tubes. Genotypes were re-confirmed post hoc by tail-PCR.Go/No-Go vibrotactile decision-making task Setup The vibrotactile decision-making setup was positioned in an isolation cubicle to minimize interference during the experiment. Mice were placed in a body tube and were head-fixed with their forepaws resting on two steel bars (6 mm diameter, Thorlabs). The right bar was mounted to a Preloaded Piezo Actuator equipped with a strain gauge feedback sensor and controlled (P-841.6 and E-501, Physik Instrumente) in a closed loop. Water reward was delivered through a metal feeding needle (20G, 1,9mm tip, Agntho's AB) connected to a lickport interface with a solenoid valve (Sanworks) equipped with a capacitive sensor (https://github.com/poulet-lab/Bpod_CapacitivePortInterface). The behavioral setup was controlled by Bpod (Sanworks) through scripts in Python (PyBpod, https://pybpod.readthedocs.io/en/latest/). Go/No-Go vibrotactile task training and testing Habituated 8-week-old mice were trained to associate a vibrotactile stimulus (pure sinusoid, 500 ms duration, 15 µm amplitude, 40 Hz frequency) with a water reward (8 µl). Go trials consisted of stimulus delivery followed by a 2 s response window during which the mice could lick to receive the reward. No-Go (catch) trials had no stimulus, and licking triggered a 5 s timeout. Inter-trial intervals varied between 5-10 s. Training included three phases: (a) Automatic water delivery at response window start. (b) Pre-training: lick-triggered water delivery on Go trials and timeout on No-Go trials. (c) Training: lick-triggered reward only if mice refrained from licking during the 3–8 s inter-trial interval before stimulus delivery; licking during this interval or on No-Go trials resulted in a 5 s timeout. Sessions consisted of 300 pseudorandomized trials: 70–80% Go, 20–30% No-Go. Pilot experiments with an extra sensor to monitor forepaw placement confirmed that the mice did not remove their forepaws from the bar before stimulus delivery. Pre-training criteria: ≥80% successful Go trials, Two-photon calcium imagingCalcium measurements were performed using a Femtonics FEMTOSmart microscope coupled with a widely tunable, femtosecond Ti:Sapphire Laser (Chameleon Ultra, Coherent) pumped by a solid-state laser (Verdi 18W, Coherent). Excitation at 920 nm was used for GCaMP8m and at 1000 nm for mRuby3 and two photomultiplier tubes were used for the collection of green and red fluorescence. Laser power was modulated using a Pockels cells A 20x/1 NA objective (Zeiss) was used to obtain 512 × 512-pixel images covering 480 × 480 μm of cortex were acquired at 30.96 Hz using the resonant scanner system (Femtonics). Measurements were written as a series of 32-bit images and saved as ome files. Analog and digital inputs sent by Bpod and the piezoelectric sensor controller were collected and saved as timestamps by the microscope and were used to synchronize the behavioral events with the calcium traces from imaging.