Background: Early childhood educators (ECEs) are the primary daytime role models for many young children, and are responsible for facilitating physical activity (PA) opportunities and minimizing sedentary behaviour (SB) in childcare. However, they have reportedly received little related education in their pre-service training. The purpose of the Training pre-service EArly CHildhood educators in physical activity (TEACH) pilot study was to explore changes in pre- and in-service ECEs’ knowledge, self-efficacy, behavioural intention, and perceived behavioural control following the TEACH e-Learning course in PA and SB. Methods: Pre-service ECEs were purposefully recruited from three Canadian colleges, while in-service ECEs were recruited via social media. A pre-post study design was used. ECEs completed two online surveys; one prior to, and one immediately following the completion of the TEACH e-Learning course (~ 5 h). Descriptive statistics were reported, and McNemar Chi-Square tests and paired samples t-tests were used to examine changes in ECEs’ question-specific, and total knowledge scores, respectively. Wilcoxon Signed Ranks tests were employed to examine changes in self-efficacy, behavioural intention, and perceived behavioural control. Results: Both pre- (n = 32) and in-service (n = 121) ECEs significantly increased their total knowledge scores from pre- to post-course completion (p <.05*). Significant positive changes in self-efficacy (p <.025*), behavioural intention (p <.007*), and perceived behavioural control (p <.007*) were demonstrated by in-service ECEs following course completion, while only select composite scores within these tools were significant among pre-service ECEs. Conclusions: These findings provide preliminary evidence of the potential efficacy of the e-Learning course at improving ECEs’ knowledge, self-efficacy, behavioural intention, and perceived behavioural control to support PA and minimize SB in childcare. Following the success of the pilot study, testing the effectiveness of the TEACH e-Learning course on a larger scale, with a comparison group, is warranted prior to recommending broader dissemination of the training in pre-service ECE programs and for in-service ECE professional learning.