The plasma membrane contains distinct domains that are characterized by a high concentration of sphingolipids and cholesterol. These membrane microdomains also referred to as rafts, seem to be intimately involved in transmembranous signaling and often initiate interactions of pathogens and the host cell membranes. Here, we investigated the further reorganization of membrane rafts in cultured epithelial cells and ex vivo isolated nasal cells after infection with rhinoviruses. We demonstrate the formation of ceramide-enriched membrane platforms and large glycosphingolipid-enriched membrane domains and the co-localization of fluorochrome-labeled rhinoviruses with these membrane domains during attachment and uptake of human rhinovirus. Destruction of glycosphingolipid-enriched membrane domains blocked infection of human cells with rhinovirus. Furthermore, our studies indicate that the activation of the acid sphingomyelinase (ASM) is intrigued in the formation of ceramide- or GM1- enriched membrane platforms. Inhibition of the ASM reduces the number of ceramide-enriched platforms and glycosphingolipid-enriched membrane domains. These data reveal a critical role of the ASM for the formation of membrane platforms and infection of human cells with rhinoviruses.