Reproduction strongly influences metabolism, morphology and behavior in female mammals. In species in which males provide parental care, reproduction might have similar effects on fathers. We examined effects of an environmental challenge on metabolically important physiological, morphological and behavioral measures, and determined whether these effects differed between reproductive and non-reproductive males in the biparental California mouse (Peromyscus californicus). Males were paired with an ovary-intact female, an ovariectomized female treated with estrogen and progesterone to induce estrus, or an untreated ovariectomized female. Within each group, half of the animals were housed under standard laboratory conditions and half in cages requiring them to climb wire towers to obtain food and water; these latter animals were also fasted for 24 h every third day. We predicted that few differences would be observed between fathers and non-reproductive males under standard conditions, but that fathers would be in poorer condition than non-reproductive males under challenging conditions. Body and fat mass showed a housing condition×reproductive group interaction: the challenge condition increased body and fat mass in both groups of non-reproductive males, but breeding males were unaffected. Males housed under the physical and energetic challenge had higher blood lipid content, lower maximal aerobic capacity and related traits (hematocrit and relative triceps surae mass), increased pain sensitivity and increased number of fecal boli excreted during tail-suspension tests (a measure of anxiety), compared with controls. Thus, our physical and energetic challenge paradigm altered metabolism, morphology and behavior, but these effects were largely unaffected by reproductive condition.