Alcohol has a variety of short‐ and long‐term effects on cell‐mediated and humoral immune response. Herein, we have characterized the impact of high‐dose EtOH administration on phenotypic and functional features of murine APC subsets, including dendritic cell (DC), macrophages and B cells. Impaired cytokine synthesis and Leishmania‐phagocytosis was observed in peritoneal macrophages following EtOH administration. Moreover, EtOH exposure led to decreased levels of splenic myeloid DC and increased percentage of macrophages with no changes in splenic lymphoid DC and B cells. Adverse effects of short‐term EtOH administration also resulted in impaired OVA‐endocytosis by DC and macrophages. In contrast, EtOH consumption upregulates OVA‐internalization by B cells. These changes on APC hierarchy may play a role shifting the fate of the immune response after EtOH ingestion. In addition to an overall downregulation of Toll‐like receptor‐TLR‐4 expression by splenic APC, a downregulation of TLR‐2 expression in macrophages was observed. Moreover, EtOH exposure altered the expression of co‐signalling molecules on splenic APC, downregulating CD40 on macrophages and upregulating CD80 on B cells, with no impact on DC subsets. The net result of changes in TLR‐mediated and co‐stimulatory signals may determine the altered immunological status induced by acute consumption of alcohol. A direct impact of high‐dose EtOH administration in the activation status of splenic CD4+ T cells was observed. Together, our results demonstrated that short‐term high‐dose EtOH administration has differential impact on APC populations, downregulating splenic macrophages and DC activity but up‐regulating B lymphocyte function as APC, and ultimately yielding a micro‐environment that led to increased activation of CD4+ T cells.