Use of cannabinoid therapies is on the rise in the United States, but responses of healthcare professionals and their knowledge of these therapies have been mixed. More information is needed about factors associated with healthcare professionals' attitudes and knowledge about medical cannabis. We conducted an online survey of US-based neurologists, nurse practitioners (NPs)/nurses, and pharmacists in August–September of 2018 (n = 451). We constructed perceived knowledge and attitudes scales and a knowledge index from multiple items and assessed state cannabis laws, participant's sociodemographics, workplace type and policies, and patient population. We used ordinary least-squares regression to examine associations among study variables. Over 80% of participants supported use and legalization of medical cannabis, especially cannabidiol (CBD) for epilepsy and when prescribed by a medical provider, but 40–50% (depending on item) felt unfamiliar with cannabinoid pharmacology and clinical applications. A total of 43% favored legal recreational cannabis. Pharmacists scored higher on the knowledge test than neurologists and NPs/nurses, but NPs/nurses had more favorable attitudes than neurologists and higher perceived knowledge than pharmacists. Both knowledge indicators predicted attitudes. State cannabis access and favorable workplace policies were associated with higher knowledge and more favorable attitudes. Healthcare professionals see potential in cannabis therapies but report significant knowledge gaps. Professional cannabinoid education is needed to address growing patient and provider demand for knowledge about cannabinoid therapies.