Figure 8. Cbln1 KO causes a major impairment in motor behaviors that is aggravated by additional deletion of Cbln2 even though the Cbln2 KO alone produces no phenotype. A, Cbln1 KO, Cbln1/2 dKO, and Cbln1/2/4 tKO impair postural sway, measured as the area around the center of force of a mouse monitored on a force plate actometer at three successive locations. Experiments were performed on two separate mouse cohorts as follows: left graph, Cbln1 KO (n = 8), Cbln2 KO (n = 9), Cbln1/2 dKO (n = 13), and control mice (Ctl; n = 10); right graph, Cbln1/2 dKO (n = 14), Cbln1/2/4 tKO (n = 11), and Ctl mice (Ctl.; n = 16). Postural sway impairments in Cbln1 KO mice were enhanced in Cbln1/2 dKO mice (adjusted p = 0.0418, Tukey's post hoc test), whereas Cbln2 KO mice exhibited no phenotype and Cbln1/2/4 tKO mice did not significantly differ from Cbln1/2 dKO mice. +p < 0.05, ****p < 0.0001, one-way ANOVA. B, Cbln1/2 dKO causes a loss of balance in mice, resulting in lateral falls, that is more severe than that observed in Cbln1 KO or Cbln1/2/4 tKO mice. A lateral fall was counted when all 4 paws were off the ground and the mouse was laying on its left or right side; the number of lateral falls during a 5 min open-field test was measured at different time points. Left, Lateral falls were observed in the Cbln1/2 dKO (n = 11) mice beginning at ∼2 months of age and the frequency of falling increased as the mouse aged. Some Cbln1 KO mice (n = 7) exhibited this loss of balance starting at 6 months of age, but the phenotype emerged earlier and was more severe in Cbln1/2 dKO mice (time *p = 0.0458; genotype ****p < 0.0001; interaction *p = 0.0385, two-way ANOVA). No lateral falls were ever observed in Ctl or Cbln2 KO mice. Right, Surprisingly, Cbln1/2/4 tKO mice (n = 11) exhibited significantly fewer lateral falls than Cbln1/2 dKO mice across all time points (n = 13; time ****p < 0.0001, genotype ****p < 0.0001, interaction *p = 0.0161, two-way ANOVA). C, Distance traveled across an 80-cm-long balance beam (width = 2 or 1 cm) by 3-month-old mice of the indicated genotypes. Left, On the 2 cm beam, the performance of Cbln1KO (n = 8) and dKO (n = 13) mice did not differ significantly from each other, but on the 1 cm beam, dKO mice traveled less distance (before falling off the beam) than Cbln1KO mice (adjusted p = 0.0005, Tukey's post hoc test, denoted by ***); both groups traveled significantly less distance compared with Ctl (n = 10) and Cbln2KO (n = 9) mice on the 2 cm (p = 0.0003, one-way ANOVA, denoted by +++) and 1 cm (p < 0.0001, one-way ANOVA, denoted by ****) beams. Right, Cbln1/2 dKO (n = 13) and Cbln1/2/4 tKO mice (n = 11) exhibited decreased performance on the balance beam as a function of age, with a dramatic decrease in the traversed distance compared with Ctl (n = 16) seen at 3 and 4 months. ****p < 0.0001, two-way ANOVA. D, Cbln1 KO, Cbln1/2 dKO, and Cbln1/2/4 tKO equally impair motor coordination and motor learning as monitored on the accelerating rotarod, whereas the Cbln2 KO has no effect. Left, Comparison of Cbln1 KO (n = 7), Cbln2 KO (n = 9), Cbln1/2 dKO (n = 13) and Ctl (n = 10) mice. Right, Cbln1/2 dKO mice (n = 14) and Cbln1/2/4 tKO mice (n = 11) exhibit similar impairments in motor coordination and motor learning compared with Ctl mice (n = 16). ****p < 0.0001, two-way ANOVA.