All tissues are genetically programmed to acquire an optimal size that is defined by total cell number and individual cellular dimensions. The retina contains stereotyped proportions of one glial and six neuronal cell types that are generated in overlapping waves. How multipotent retinal progenitors know when to switch from making one cell type to the next so that appropriate numbers of each cell type are generated is poorly understood. Pten is a phosphatase that controls progenitor cell proliferation and differentiation in several lineages. Here, using a conditional loss-of-function strategy, we found thatPtenregulates retinal cell division and is required to produce the full complement of rod photoreceptors and amacrine cells in mouse. We focused on amacrine cell number control, identifying three downstream Pten effector pathways. First, phosphoinositide 3-kinase/Akt signaling is hyperactivated inPtenconditional knock-out (cKO) retinas, and misexpression of constitutively active Akt (Akt-CA) in retinal explants phenocopies the reduction in amacrine cell production observed inPtencKOs. Second, Akt-CA activates Tgfβ signaling in retinal explants, which is a negative feedback pathway for amacrine cell production. Accordingly, Tgfβ signaling is elevated inPtencKO retinas, and epistatic analyses placed Pten downstream of TgfβRII in amacrine cell number control. Finally,Ptenregulates Raf/Mek/Erk signaling levels to promote the differentiation of all amacrine cell subtypes, which are each reduced in number inPtencKOs.Ptenis thus a positive regulator of amacrine cell production, acting via multiple downstream pathways, highlighting its diverse actions as a mediator of cell number control.SIGNIFICANCE STATEMENTDespite the importance of size for optimal organ function, how individual cell types are generated in correct proportions is poorly understood. There are several ways to control cell number, including readouts of organ function (e.g., secreted hormones reach functional levels when enough cells are made) or counting of cell divisions or cell number. The latter applies to the retina, where cell number is regulated by negative feedback signals, which arrest differentiation of particular cell types at threshold levels. Herein, we show thatPtenis a critical regulator of amacrine cell number in the retina, acting via multiple downstream pathways. Our studies provide molecular insights into howPTENloss in humans may lead to uncontrolled cell division in several pathological conditions.