A one-pot access to the protoberberine and tetrahydroprotoberberine scaffold has been developed from easily available primary amine-functionalized o-alkynylbenzaldehydes. The core skeleton of protoberberine was afforded via in situ generation of isochromenylium by Ag(I) catalysis, subsequent cyclization to isoquinolinium, and final hydride reduction by a continuous operation procedure. The newly developed step-economic protocol shows excellent regio- and stereoselectivity and is capable of achieving those previously unavailable derivatives/analogues with electron-deficient substituents, providing a robust tool for medicinal investigations of protoberberines and derivatives.