(A–D) Facial motor nucleus cryosections from neonatal wild-type (A and B) and Etv1 mutant (C and D) mice showing retrograde Rh-Dx labeling from NL and OO motor fibers (red) overlaid with Alcam expression (blue). Boxes denote positions of high-power images (A’–D’). Selectivity of Alcam expression in NL (A’, arrows) and its exclusion from OO (B’, arrowheads) FMNs is lost in Etv1 mutants, with many AlcamOFF FMNs innervating the NL (arrowheads in C’) and AlcamON FMNs innervating the OO (arrows in D’). Arrow head in (A’) marks a likely AlcamOFF NL FMN.
(E) Coincidence of Alcam NL pool-marker expression in OO or NL FMNs, as represented by the percentage (± SE) of FMNs marked by retrograde muscle labeling that express Alcam. The specificity of NL innervation by AlcamON FMNs in control mice is lost in Etv1 mutants. FMNs marked by OO and NL retrograde labeling were scored for colocalization of Alcam in control and age-matched Etv1nlz/nlz mice. Data represented as the percentage (± SE) of FMNs marked by retrograde label and Alcam expression, n = 3 mice per genotype, 52 NL and 47 OO FMNs from control mice and 98 NL and 47 OO FMNs from Etv1-mutant mice (unpaired t test, *p < 0.05, **p < 0.0005). Scale bar: 200 μm.
(F–I) Eyelid behavior in control (F and G) and Etv1-mutant (H and I) mice at P21 before (F and H) and after (G and I) Z/T facial nerve-branch axotomy. OO axotomy partially rescued tonic eyelid closure in Etv1 mutants, whereas the dimensions of the palpebral fissure remained unchanged in controls.
(J) Quantitation of axotomy-induced changes in palpebral fissure dimensions. Values represented as PCI means ± SE, n = 4 mice per genotype (unpaired t test, *p < 0.001, **p = 0.0001).
(K–M) Electromyographic (EMG) recordings from the OO and NL muscles of control (K and L) and Etv1-mutant (M) mice at P20. OO muscle did not contract during exploratory whisking in control mice (K) but in Etv1-mutant mice, the OO was activated simultaneously and synchronously with the NL during bouts of exploratory whisking (M). n = 8 control, 6 mutant mice.
(N) Diagram of changes in peripheral facial-motor circuitry leading to OO/NL synkinesis in Etv1 mutants.
See also Figure S7.