To determine whether retinal degeneration exacerbated the behavioral phenotypes of mice with circadian mutations, we observed the wheel running behavior of mice lacking
mCry genes,
mPer genes, or
Bmal1 with and without
rd/rd. Representative actograms from mice kept in a 12-hour light/12-hour dark cycle with a phase shift of the light-dark cycle and the dark-dark cycle are shown in
Figure 5. Wild-type and
rd/rd mice typically demonstrated entrainment with nocturnal phase preference in 12-hour light/12-hour dark, gradual re-entrainment to an altered phase of 12-hour light/12-hour dark, and free-running rhythmicity in dark-dark with a period of approximately 23.6 hours (
Figs. 5A,
5B). In contrast, cryptochrome mutant mice demonstrated an immediate shifting of phase with a new light cycle, as previously reported
27 (
Fig. 5C). In contrast to the strong masking behavior seen in cryptochrome mutant mice,
Period and
Bmal mutant mice both demonstrated relatively weak rhythmicity in light-dark conditions (
Figs. 5E,
5G). When kept in constant darkness,
mCry1 −/− ;mCry2 −/− ; mPer1 −/− ;mPer2 −/−, and
Bmal1 −/− mice are behaviorally arrhythmic.
37,38 In general
Bmal1 −/− exhibited the least overall activity and the weakest rhythmicity, with one third of the animals showing no rhythmic behavior in any lighting condition.
Period mutant mice had an intermediate phenotype exhibiting moderate overall levels of activity and some visible rhythmicity in every animal. In all cases, however, the three clock mutant phenotypes were indistinguishably arrhythmic in dark-dark.
The addition of outer retinal degeneration to clock mutation induced more arrhythmicity in light-dark conditions in some animals. With the addition of rd/rd, the robustness of masking in some cryptochrome mutants decreased as activity became less consolidated to dark hours. A subset of mPer1 −/− ;mPer2 −/− ;rd/rd and Bmal1 −/− ;rd/rd became arrhythmic in all lighting conditions, whereas some mice retained some weak rhythmicity. With the addition of retinal degeneration, some Bmal1 −/− mice exhibited some rhythmicity in light-dark only but with a strange phase relationship; 2 of 6 mice had weak rhythmicity in 12-hour light/12-hour dark, with activity onset at 1 hour after lights off, and two other mice exhibited a diurnal preference but with activity onset delayed to the middle of the light phase. A subset of Bmal1 −/− ;rd/rd mice exhibited arrhythmicity in all lighting conditions.