Validation of the CD11c‐DTR transgenic mouse as a model to study neuroimmune interactions in the cornea

Purpose: CD11c‐DTR+ mice have a transgene encoding for a simian diphtheria toxin (DT) receptor under the CD11c promoter that makes dendritic cells (DCs) sensitive to DT, which causes DC depletion. Preliminary results using this mouse suggest that there is a functional interaction between DCs and corneal sensory nerves, as DC depletion modifies nerve activity and nociceptive behaviour. The aim of this work was to validate the CD11c‐DTR+ mouse model proving that the observed effects are not due to the DT itself, using littermates negative for the transgene and therefore resistant to DC depletion by DT. Methods: CD11c‐DTR+ and CD11c‐DTR‐ mice (5 months, both sexes) were used. Both groups received single or repetitive (every 2 days for 8 days) subconjunctival injections of DT or its vehicle (PBS). Basal tearing (in mm) was measured using phenol red threads placed in the temporally side of the eye for 30 s in isoflurane‐anaesthetised mice. To quantify spontaneous eye pain, the eye closure ratio was calculated by dividing the height by the width of the lid opening measured in photographs of freely moving animals. Nerve terminal impulse (NTI) activity of corneal cold thermoreceptor endings was recorded ex vivo in superfused eyes; spontaneous and stimulus‐evoked activity was explored. Results: Basal tearing was higher in DT‐injected CD11c‐DTR‐ animals, especially after repetitive injections (Day 0: 2 ± 0.16 mm vs. day 2: 2.8 ± 0.37 mm vs. day 8: 4.7 ± 0.36 mm, n = 5, p ≤ 0.001, RM ANOVA). Repetitive PBS injections caused similar effects in CD11c‐DTR+ and CD11c‐DTR‐ mice tearing (DTR−: 4.7 ± 0.36 mm, n = 5 vs. DTR+: 3.7 ± 1.48 mm, n = 3, p = 0.399, t‐test). No signs of spontaneous pain were observed in DT‐injected CD11c‐DTR‐ mice (Day 0: 0.91 ± 0.01 vs. 48 h: 0.93 ± 0.01 vs. day 8: 0.95 ± 0.01, n = 5, p = 0.065, RM ANOVA). No significant differences were found in spontaneous and stimulus‐evoked NTI activity recorded after repetitive injections of DT in CD11c‐DTR− mice or PBS in CD11c‐DTR+ mice. Conclusions: CD11c‐DTR+ mice are a valid model to study neuroimmune interactions. DT has no effects on pain behaviour or NTI activity of animals lacking the DT receptor. The changes in tearing appear to be consequence of the subconjunctival injection process per se. Reference PID2020‐115934RB‐I00 from DOI: MCIN/AEI/10.13039/50110001103