Low back pain (LBP) is a highly prevalent and disabling condition whose initiating factors are poorly understood. It is known that psychological and physical stress is associated with LBP but the causal relationship, mechanisms, and mediators have not been elucidated, and a preclinical model enabling the investigation of causality and thereby critically contributing to clinical translation does not exist. In this study, we first established and characterized a myofascial LBP model in mice based on nerve growth factor (NGF) injection into the low back muscles. Second, we investigated the effect of 2 different stress paradigms on this mouse LBP model by applying the chronic unpredictable stress and vertical chronic restraint stress (vCRS) paradigms, to mimic psychological and psychophysical stress, respectively. In these studies, we combined longitudinal behavioral tests with gene and protein expression analysis in the muscle, dorsal root ganglia, and spinal cord. Nerve growth factor-induced LBP was characterized by long-lasting local and plantar mechanical hypersensitivity, cold hyperalgesia, decreased grip strength and wheel running activity, and time-dependent changes of neuropeptide and glial markers in the spinal cord. Interestingly, the exposure to chronic unpredictable stress slightly worsened pain behavior, whereas vCRS primed and highly aggravated pain in this LBP model, by causing per se the intramuscular upregulation of endogenous NGF and increased spinal astrocyte expression. Our mouse model, particularly the combination of NGF injection and vCRS, suggests that similar mechanisms are important in nonspecific LBP and might help to investigate certain aspects of stress-induced exacerbation of pain.