Previous studies suggest that melanocortin-4 receptor (MC4R) activation in areas outside of the hypothalamus may regulate appetite but the role of MC4R in specific neuronal populations of the hindbrain in regulating cardiovascular function are still unknown. We examined the impact of total body MC4R deficiency (LoxTB-MC4R mice) and restoration of MC4R specifically in the brainstem and/or spinal cord on blood pressure (BP) response to acute stress. We selectively rescued MC4R only in preganglionic parasympathetic neurons of the dorsal motor of the vagus (DMV) and in the nucleus tractus solitarius (NTS) (LoxTB-MC4R/Phox2B-cre mice, n=5) or in cholinergic preganglionic neurons of the hindbrain and spinal cord (LoxTB-MC4R/Chat-cre mice, n=4). Mice were implanted with telemetry probes for measurement of mean arterial pressure (MAP) and heart rate (HR). After a 10-day recovery period, MAP and HR were continuously measured for 30 minutes before, during and 30 minutes after an air jet stress test. Acute air jet stress significantly increased MAP by 33±3 in WT mice and 31±2 in LoxTB-MC4R/Chat-cre mice compared to only 18±4 or 20±3 mmHg in LoxTB-MC4R/Phox2B-cre mice and LoxTB-MC4R mice, respectively. HR was increased by 180±20, 110±20, 100±23, and 127±15 bpm in response to air jet stress in WT, LoxTB-MC4R, LoxTB-MC4R/Phox2B and LoxTB-MC4R/Chat-cre mice, respectively. These results indicate that MC4Rs in cholinergic preganglionic neurons of the hindbrain and spinal cord play a key role in the BP responses to acute stress. (NHLBI-PO1HL51971, NIGMS- P20GM104357, and AHA-SDG5680016)