The plant growth regulator Mepiquat Chloride (MC) is widely used in cotton (Gossypium hirsutum L.) production to control plant vegetative growth, and in many cases, results in increased yields. Since plant growth response is highly dependent on temperature, it has been speculated that inconsistent responses to MC may be related to the temperature under which the plant is growing at the time of application. It has been reported that in extreme temperature conditions, the effect of MC was either decreased or very variable, with inconclusive explanations. The objective of this study was to determine the critical concentration of MC in cotton plants as affected by temperature regimes. Cotton plants were grown in growth chamber conditions and, at pinhead-square, MC was applied at 15.0 and 30.0 g ha-1 of active ingredient. Plants were then, submitted to three temperature regimes: 25/15 oC, 32/22 oC, and 39/29 oC (day/night temperatures). A treatment without MC application was used as a control. The results showed that the largest effect of the plant growth regulator on cotton growth occurred when the daily temperature was 32 °C, with nights of 22 °C. This temperature regime resulted in the optimum growth response to MC with better distinction of the growth curves based on the rates of growth regulator applied.