The objective of this trial was to evaluate the effects of feed restriction (FR) on serum glucose, nonesterified fatty acids, progesterone (P4), insulin, and milk production in dairy cows. Eight multiparous Holstein cows, 114±14 d pregnant and 685±39kg of body weight, were randomly assigned to a replicated 4×4 Latin square design with 14-d periods. During the first 8 d of each period, cows in all treatments were fed for ad libitum feed intake. Beginning on d 9 of each period, cows received 1 of 4 treatments: ad libitum (AL), 25% feed restriction (25FR), 50% feed restriction (50FR), and 50% of TMR replaced with wheat straw (50ST). Daily feed allowance was divided into 3 equal portions allocated every 8h with jugular blood samples collected immediately before each feeding through d 14. In addition, on d 12 of each period, blood samples were collected before and at 60, 120, 180, 240, 300, 360, 420, and 480min after morning feeding. The conventional total mixed ration and total mixed ration with straw averaged 15.1 and 10.8%, 32.1 and 50.5%, and 26.8 and 17.0% for concentrations of crude protein, neutral detergent fiber, and starch, respectively. Cows that were feed and energy restricted had reduced dry matter intake, net energy for lactation intake, circulating glucose concentrations, and milk production, but greater body weight and body condition score losses than AL cows. Circulating concentrations of insulin were lower for cows fed 50FR (8.27μIU/mL) and 50ST (6.24μIU/mL) compared with cows fed AL (16.65μIU/mL) and 25FR (11.16μIU/mL). Furthermore, the greatest plasma nonesterified fatty acids concentration was observed for 50ST (647.7μEq/L), followed by 50FR (357.5μEq/L), 25FR (225.3μEq/L), and AL (156.3μEq/L). In addition, serum P4 concentration was lower for cows fed AL than cows fed 50ST and 25FR. Thus, FR reduced circulating glucose and insulin but increased P4 concentration, changes that may be positive in reproductive management programs.