The present disclosure provides display substrate and a fabrication method thereof, a display panel and a display device. The display substrate has a plurality of pixel regions, and includes an anode, a light-emitting layer and a cathode in each pixel region. In each pixel region, the light-emitting layer includes a plurality of light-emitting units spaced apart from each other, both the anode and the cathode are conductive layers each having an integral structure, and the plurality of light-emitting units are connected to the anode and the cathode, respectively.