Three experiments were conducted (1) to evaluate the effects of supplementing a reduced-protein diet with a ruminally available source of N and rumen-protected (RP) limiting AA on performance of lactating cows; (2) to determine the specific effects of His supplementation on lactation performance of dairy cows fed a reduced-protein diet; and (3) to investigate the effects of feeding a His-deficient diet on lactational performance and the endogenous His-reserves of dairy cows. The objective for the first study was to investigate the effects of slow-release urea and rumen-protected Met (RPMet) and rumen-protected His (RPHis) supplementation of a reduced-protein diet [deficient in metabolizable protein (MP), according to the National Research Council (NRC, 2001)] on lactation performance of dairy cows. We hypothesized that slow-release urea supplementation would alleviate the previously observed negative effect of lower ruminal N availability on total-tract fiber digestibility. We also hypothesized that supplementation of RPMet and specifically RPHis may increase dry matter intake (DMI), milk yield, and milk protein yield (MPY). Sixty lactating Holstein cows were used in a 10-wk randomized complete block-design trial. Cows were fed a covariate diet for 2 wk (i.e., baseline period during which all cows are fed the same diet) and then assigned to one of the following treatments: MP-adequate diet (AMP), MP-deficient diet (DMP), DMP supplemented with slow-release urea (DMPU), DMPU supplemented with RPMet (DMPUM), and DMPUM supplemented with RPHis (DMPUMH). The DMP diet did not affect DMI, yields of milk, and milk components, despite a reduction in nutrient digestibility, compared to AMP. Urinary N and urea-N excretions were decreased by DMP, compared to AMP. Cows fed DMP had higher plasma concentration of 3-methylhistidine and gained less body weight (BW) compared to AMP and cows fed DMPU, DMPUM, and DMPUMH. Addition of slow-release urea to the DMP diet increased urinary urea-N excretion. Supplementation of RPMet increased plasma Met concentration but had no effect on milk production or composition. The DMPUMH increased DMI, milk true protein concentration and yield, and decreased milk fat concentration, compared to DMPUM. These results were in line with a previous study from A. N. Hristovs laboratory and suggested that His may have a positive effect on feed intake and milk production and composition in dairy cows fed MP-deficient diets.The objective of the second experiment was to evaluate the effects of supplementing a reduced-protein diet [deficient in MP, based on NRC (2001)] with RPMet, rumen-protected Lys (RPLys), and RPHis, individually or combined, on the performance of lactating dairy cows. We hypothesized that supplementation of (1) RPHis may improve DMI, milk yield, and MPY; (2) RPMet, RPLys, or both may increase milk protein content and MPY; and (3) a combination of the 3 RPAA (Met, Lys, His) may further increase milk yield and MPY. The experiment was a 9-wk randomized complete block design with 72 Holstein cows. Following a 2-wk covariate period, cows were assigned to 1 of the following 6 treatments: MP-adequate diet (MPA), MP-deficient diet (MPD), MPD supplemented with RPMet (MPDM); (4) MPD supplemented with RPLys (MPDL); (5) MPD supplemented with RPHis (MPDH); and (6) MPD supplemented with RPMet, RPLys, and RPHis (MPDMLH). The MPD diet decreased DMI, yields of milk and milk components (fat, protein, lactose), energy-corrected milk (ECM), and feed efficiency, compared to MPA. Cows fed MPD had lower milk and plasma urea N and higher milk N efficiency compared to cows fed MPA. Supplementation of MPD with RPLys increased milk protein content. Addition of RPHis increased DMI and milk protein concentration. Supplementation of the 3 RPAA increased yields of milk fat, protein, and ECM and ECM feed efficiency. Cows fed MPD had lower blood hemoglobin (Hb) concentration than cows fed MPA. Overall, data from this study confirmed our previous findings and suggested that His stimulates DMI and the combination of the 3 RPAA (Met, Lys, and His) has the potential to improve milk and milk component yields in dairy cows fed MP-deficient diets.The objective of the third experiment was to investigate the effects of feeding a His-deficient diet on lactational performance and the endogenous His-reserves of dairy cows. We hypothesized that feeding a His-deficient diet may decrease DMI, yields of milk and/or milk components, BW gain, N efficiency, and the endogenous His-reserves (i.e., blood Hb and muscle His-dipeptides). Supplementation of the His-deficient diet with RPHis may alleviate these effects in dairy cows. The study was a 10-wk randomized complete block design with 24 Holstein cows. After a 2-wk covariate period, cows were assigned to a His-adequate diet (HAD) and His-deficient diet (HDD). At the end of the 10-wk experiment, HDD was supplemented with RPHis for an additional 9 d. The HDD diet decreased DMI, yields of milk, protein and lactose, ECM, and milk and plasma urea N, compared to HAD. Total-tract apparent digestibility of dry and organic matter, crude protein, and neutral detergent fiber, and excretion of urinary N and urea N were decreased by HDD, compared to HAD. Concentrations of muscle His, plasma His and blood Hb were decreased by HDD compared to HAD, suggesting a supply of His from the endogenous reserves. The 9-d supplementation of HDD with RPHis increased DMI and ECM yield, compared to HDD. These results confirmed the findings of the two previous experiments that low dietary His supply impair DMI and yields of milk and milk protein in dairy cows. Supplementation of such diets with RPHis has the potential to reverse these effects.