Drug developers worldwide assess compound safety and efficacy using measures that include mouse core temperature and locomotor activity. Subtle differences in animal housing conditions between institutions can alter these values, impacting scientific rigor and reproducibility. In these studies, adult male NIH Swiss mice were surgically implanted with radiotelemetry probes that simultaneously monitored core temperature and locomotor activity across various housing conditions. In the first study, ambient temperature was varied between 20 °C and 28°C in groups of singly housed mice. Additional studies held the mice at a constant ambient temperature and examined the effects of cage density (housing animals singly or in groups of 3 or 6), bedding change and provision of nesting material, and the availability of a running wheel on core temperature and locomotor activity. Mice overwhelmingly maintained species-typical core temperatures across all ambient temperatures, across all housing conditions, when bedding was fresh or old, and with or without the provision of cotton squares as nesting material. However, engaging in wheel running and the combination of fresh bedding and cotton squares transiently increased core temperatures beyond the species-typical range. Similarly, the circadian distribution of locomotor activity was significantly disrupted by placing animals in cages with fresh bedding or nesting material, or by performing both of these manipulations concurrently during the light period. These findings suggest that standard husbandry practices and common housing conditions may transiently affect core temperature in adult mice. Furthermore, these practices may have profound and relatively long-lasting effects on motor activity and the regulation of circadian rhythms.