The effects of subcutaneous injection of glucagon and/or oral administration of glycerol on blood metabolites and hormones of Holstein dairy cows induced with fatty liver disease was studied. Twenty multiparous cows were fed a dry cow ration supplemented with 6 kg of cracked corn during the dry period to increase the likelihood of fatty liver disease development. Cows with a body condition score of ≥ 3.5 points (1-5 scale) were assigned randomly to one of four treatment groups--saline, glucagon, glucagon plus glycerol, and glycerol. Following treatment, serial blood samples were collected to determine the effect of glucagon and/or glycerol on blood composition. Glucagon injection alone increased postpartal plasma glucose, insulin, glucagon, and plasma urea nitrogen and decreased plasma nonesterified fatty acids (NEFA). Glucagon plus glycerol treatment increased and sustained postpartal plasma glucose, insulin and decreased postpartal plasma NEFA. Administration of glycerol alone increased plasma triacylglycerol (TAG) and decreased plasma NEFA during the postpartal period. Early postpartal treatment of cows with glucagon and/or glycerol increased plasma glucose and decreased plasma NEFA. This response would suggest that these treatments with glucagon and/or glycerol would decrease the likelihood of fatty liver disease in dairy cows.