Gap junction channels formed by the association of connexin hemichannels play a crucial role in intercellular communication. Connexin 43 (Cx43) is expressed in a variety of tissues and organs, including heart and brain, and abnormal sustained opening of undocked "free" hemichannels contributes to the cell damage in cardiac infarcts and stroke. Selective inhibitors of Cx43 hemichannels for clinical use are then desirable. Here, we synthesized and tested new aminoglycosides for their connexin inhibitory activity towards Cx26 and Cx43 hemichannels. The lead compounds displayed enhanced Cx43/Cx26 selectivity for hemichannel inhibition when compared to the parent kanamycin A and other commercially available aminoglycosides. These lead compounds are not cytotoxic to mammalian cells and show promise for the treatment of ischemic damage of the heart, brain, and kidneys. We identified a new compound as a promising lead based on its good selectivity for Cx43 hemichannels inhibition and the simplicity and affordability of its production.