To identify small molecules that can induce β-cell replication, a large chemical library was screened for proliferation of growth-arrested, reversibly immortalized mouse β cells by using an automated high-throughput screening platform. A number of structurally diverse, active compounds were identified, including phorbol esters, which likely act through protein kinase C, and a group of thiophene-pyrimidines that stimulate β-cell proliferation by activating the Wnt signaling pathway. A group of dihydropyridine (DHP) derivatives was also shown to reversibly induce β-cell replication in vitro by activating L-type calcium channels (LTCCs). Our data suggest that the LTCC agonist 2a affects the expression of genes involved in cell cycle progression and cellular proliferation. Furthermore, treatment of β cells with both LTCC agonist 2a and the Glp-1 receptor agonist Exendin-4 showed an additive effect on β-cell replication. The identification of small molecules that induce β-cell proliferation suggests that it may be possible to reversibly expand other quiescent cells to overcome deficits associated with degenerative and/or autoimmune diseases.