Previous research suggested that one or more choline products altered the inflammatory response of bovine immune cells supplemented with free choline. The objective was to assess the impact of choline product inhibitors on the inflammatory response of monocytes and lymphocytes from cows in early lactation (3.8 ± 2.5 DIM, n = 5). Blood immune cells were isolated using density gradient media and were incubated at 37°C and 5% CO₂. Mononuclear cells were incubated for 2 h, then monocytes were separated from lymphocytes by adherence. Monocytes (15 min) and lymphocytes (30 min) were primed with chemicals reported to inhibit the activity of choline dehydrogenase (CDI), choline kinase (CKI), muscarinic acetylcholine receptors (MRI), nicotinic acetylcholine receptors (NRI), and control without inhibitor (CTL), and then supplemented with choline (0 or 10 µM) for 1 h (monocytes) or 4 h (lymphocytes). Cells were treated with LPS (1 µg/mL, monocytes, 2 h) or concanavalin-A (5 µg/mL, lymphocytes, 48 h) to harvest spent media for cytokine measures. Another subset of monocytes (identical inhibitor and choline treatments) was treated or not with labeled E. coli for 40 min, followed by phagocytic and oxidative burst assays, and treated lymphocytes were challenged or not with concanavalin A for 48 h, followed by a 24-h proliferation assay. The data were transformed to attain normality and analyzed as a complete randomized design. Regardless of choline treatment, CDI markedly inhibited (P < 0.01) monocyte phagocytosis, oxidative burst, and TNF-α production and lymphocyte proliferation and IFN-γ production. Unlike with other inhibitors, inhibition of lymphocyte proliferation by CDI was reversed (P < 0.01) by activation with concanavalin A (~3 ×). Supplemental choline enhanced (P < 0.01) phagocytosis and oxidative burst of CTL- and CKI-treated monocytes but reduced that of MRI-treated monocytes. These findings confirm that choline affects the inflammatory response of innate and adaptive immune cells via its downstream products, contributing to the efforts to define or refine choline requirements in transition cows.