The Holocene Megathermal is divided into early, middle and late periods, each having different impacts on the Neolithic cultures due to their different climate changing trends. This study is based on a comparative analysis of the environmental evolution information recorded in the Qinghai Lake, the western edge of the Loess Plateau and Zoige and the spatial distribution of Neolithic sites of the Gansu-Qinghai region. Results show that the early and middle periods towards warm and humid promoted the development of Neolithic cultures with agriculture as the main sector in the Gansu-Qinghai region, furthermore a heyday of Yangshao Culture prosperity emerged. The Holocene entered the late period after the cold climate event at 5.9 ka BP in the Gansu-Qinghai region. Later the climate began to turn cold. However, at the 5.8–4.2 ka BP a relatively stable warm and humid climate created the conditions for the development of Majiayao Culture of the Neolithic Age in this region, thus the distribution of its cultural heritage sites expanded towards high altitudes and high latitudes. From 4.2 ka BP onwards, the climate became cold and dry, which had a significant influence on the Neolithic cultures of the Gansu-Qinghai region, leading to a dramatic change in the cultural characters and spatial distribution of Qijia Culture around 4.2 ka BP. After a nearly 300 years of cold and dry period, the unified Neolithic farming culture completely collapsed. Afterwards an industrial division of animal husbandry and farming and regional multiple cultures formed, and ultimately led to the end of primitive society and the starting of a civilized society.