Background: Acupuncture is safe and effective for improving the motor function of poststroke hemiplegic patients, but there still exists a certain gap between clinical practice and its neural mechanisms. The cerebral functional reconstruction after unilateral motor pathway injury exhibits a bilateral tendency, however current studies seldom pay attention to it. Hence, based on bilateral cerebral connections, the underlying mechanism of acupuncture in stroke rehabilitation remains research space to date. Methods: This is a single-center, randomized controlled, paralleled neuroimaging trial, with patients, outcome assessors, and data statisticians blinded. Stroke patients with motor dysfunction meeting the inclusion criteria will be randomly assigned (2:1) to receive either true acupoints treatment or sham acupoints treatment (5 sessions per week for 2 weeks). All the patients will receive conventional treatment and rehabilitation. Participants will undergo the neuroimaging scanning before and after the entire acupuncture treatment. The clinical primary outcomes are Fugl-Meyer assessments (FMA) and the National Institutes of Health stroke scales (NIHSS). The secondary outcome is Needle Sensation Assessment Scale (NSAS). The neuroimaing outcomes are voxel-mirrored homotopic connectivity (VMHC), fractional anisotropy (FA) and gray matter volume (GMV). The correlation analysis will be explored between the neuroimage indicators and clinical motor assessments. Discussion: From the perspective of bilateral connection, this trial will investigate the cerebral influence of acupuncture treatment on poststroke patients with motor dysfunction, promoting application of acupuncture in stroke rehabilitation. Trial registration: Chinese Clinical Trials Registry, ChiCTR 1800016263. Registered on 22 May, 2018.