Microenvironment-sensitive fluorescent (ESF) nucleosides are powerful tools for nucleic acid research. The new 5-substituted uridine analogues are synthesized, which comprise a 4H-cyclopenta[d]thiazole ring obtained by the Hantzsch synthesis reaction of 5-thioamide-uridine with aromatic α-bromocarbonyl compounds. The emission maximum of new compounds is in the visible region. They exhibit strong solvent- and pH-dependent fluorescent properties, indicating their promising ability to be fluorescent probes.