Aim Skeletal muscle atrophy following prolonged immobilization (IM) is a catabolic state characterized by increased proteolysis and functional deterioration. Previous research indicates that discord of mitochondrial homoeostasis plays a critical role in muscle atrophy. We hypothesized that muscle IM would activate the ubiquitin‐proteolysis, autophagy–lysosome (mitophagy) pathway, mitochondrial dynamics remodelling and apoptosis partially controlled by the FoxO signalling pathway. Methods Female FVB/N mice were randomly divided into five groups (n = 8 each): control (CON), IM with banding of one of the hindlimbs for 1, 2 and 3 weeks (1w‐, 2w‐ and 3w‐IM) and 2w‐IM followed by 1 week of remobilization (RM). Results Mitochondrial density and DNA copies in tibialis anterior (TA) muscle were reduced by approx. 80% (P < 0.05 for 2w‐IM; P < 0.01 for 3w‐IM), along with activation of FoxO3a, atrogin‐1 and MuRF1 following 2w‐ and 3w‐IM (P < 0.01). Protein markers of autophagy/mitophagy, such as beclin 1 (approx. 2.7‐fold; P < 0.01), LC3, ubiquitin‐binding adaptor (approx. 1.47‐fold; P < 0.01), Rheb (approx. 1.9‐fold; P < 0.05) and parkin (approx. 70%; P < 0.05), were all increased by IM and remained activated after RM, whereas BNIP3 and PINK1 levels were decreased by IM (P < 0.05), but elevated upon RM (P < 0.01). IM decreased Mfn2 expression (approx. 50%; P < 0.01) and increased Fis‐1 expression (approx. 2.4‐fold; P < 0.05). Muscle apoptosis indicator Bax/Bcl2 ratio was elevated at 2w‐ to 3w‐IM (approx. 3.7‐fold; P < 0.01), whereas caspase‐3 activity was five‐ to sixfold higher (P < 0.01) and remained threefold higher above CON (P < 0.05). Conclusion Our data indicate that IM‐induced mitochondrial deterioration is associated with altered protein expressions in the autophagic/mitophagic pathway, more fragmented mitochondrial network and activation of apoptosis partly under the influence of FoxO3 activation.