Abstract Caffeine is a well described and characterized ryanodine receptor (RyR) activator. Previous evidence from independent research studies also indicate caffeine inhibits InsP 3 receptor functionality, which is important to activation of capacitative Ca 2+ entry (CCE) in some cell types. In addition, RyR activation elicits excitatory-coupled Ca 2+ entry (ECCE) in skeletal muscle myotubes. Recent studies by our group show that canine pulmonary arterial smooth muscle cells (PASMCs) have functional InsP 3 receptors as well as RyRs, and that CCE is dependent on InsP 3 receptor activity. The potential for caffeine to activate ECCE as well as inhibit InsP 3 receptor function and CCE was examined using fura-2 fluorescent imaging in canine PASMCs. The data show caffeine causes transient as well as sustained cytosolic Ca 2+ increases, though this is not due to CCE or ECCE activity as evidenced by a lack of an increase in Mn 2+ quench of fura-2. The experiments also show caffeine reversibly inhibits 5-HT elicited—InsP 3 mediated Ca 2+ responses with an IC 50 of 6.87 × 10 − 4 M and 10 mM caffeine fully inhibits CCE. These studies provide the first evidence that caffeine is an inhibitor of InsP 3 generated Ca 2+ signals and CCE in PASMCs.