A display substrate and a display device are disclosed. In the display substrate, a pixel defining layer includes a plurality of opening groups, each of the opening groups includes a first opening and a second opening, the spacer is located between the first opening of a first opening group row and the second opening of a second opening group row, an orthographic projection of the first opening includes a first long edge, and an orthographic projection of the second opening includes a second long edge, an orthographic projection of the spacer includes a third long edge, and an angle between the third long edge and the first long edge ranges from 20 degrees to 70 degrees, the orthographic projection of the first opening includes a first short edge, and the length of the third long edge is greater than that of the first short edge.