Obesity is a widespread problem across the leisure population of horses and ponies in industrialised nations. Skeletal muscle is a major contributor to whole body resting energy requirements and communicates with other tissues through the secretion of myokines into the circulation. Myostatin, a myokine and negative regulator of skeletal muscle mass, has been implicated in obesity development in other species. This study evaluated gene and protein expression of myostatin and its receptor, ActRIIB in adipose tissues and skeletal muscles and serum myostatin concentrations in six lean and six obese animals to explore putative associations between these factors and obesity in horses and ponies. Myostatin mRNA expression was increased while ActRIIB mRNA was decreased in skeletal muscles of obese animals but these differences were absent at the protein level. Myostatin mRNA was increased in crest fat of obese animals but neither myostatin nor ActRIIB proteins were detected in this tissue. Mean circulating myostatin concentrations were significantly higher in obese than in lean groups; 4.98 ng/ml (±2.71) and 9.00 ng/ml (±2.04) for the lean and obese groups, respectively. In addition, there was a significant positive association between these levels and myostatin gene expression in skeletal muscles (average R2 = 0.58; p<0.05). Together, these results provide further basis for the speculation that myostatin and its receptor may play a role in obesity in horses and ponies.