Ethnopharmacological relevance Momordica charantia fruit is a widely used traditional medicinal herb as, anti-diabetic, anti-HIV, anti-ulcer, anti-inflammatory, anti-leukemic, anti-microbial, and anti-tumor. Aims of study The present study is undertaken to investigate the possible mode of action of fruit extracts derived from Momordica charantia (MC) and study its pharmacological effects for controlling diabetic mellitus. Effects of aqueous and chloroform extracts of Momordica charantia fruit on glucose uptake and up-regulation of glucose transporter (Glut-4), peroxisome proliferator activator receptor gamma (PPARγ) and phosphatidylinositol-3 kinase (PI3K), were investigated to show its efficacy as a hypoglycaemic agent. Materials and methods Dose dependent glucose uptake assay was performed on L6 myotubes using 2-deoxy-d-[1-3H] glucose. Up-regulatory effects of the extracts on the mRNA expression level of Glut-4, PPARγ and PI3K have been studied. Results The association of Momordica charantia with the aqueous and chloroform extracts of Momordica charantia fruit at 6μg/ml has shown significant up-regulatory effect, respectively, by 3.6-, 2.8- and 3.8-fold on the battery of targets Glut-4, PPARγ and PI3K involved in glucose transport. The up-regulation of glucose uptake was comparable with insulin and rosiglitazone which was approximately 2-fold over the control. Moreover, the inhibitory effect of the cyclohexamide on Momordica charantia fruit extract mediated glucose uptake suggested the requirement of new protein synthesis for the enhanced glucose uptake. Conclusion This study demonstrated the significance of Glut-4, PPARγ and PI3K up-regulation by Momordica charantia in augmenting the glucose uptake and homeostasis.