A quantitative understanding of the bulk excitability of skeletal muscle tissues is important for the design of muscle tissue bioreactor systems, implantable muscle stimulators, and other systems where electrical pulses are employed to elicit contractions in muscle tissue both in vitro and in vivo. The purpose of the present study is to systematically compare the excitability of mammalian (rat) skeletal muscle under a range of conditions (including neonatal development, denervation, and chronic in vivo stimulation of denervated muscle) and of self-organized muscle tissue constructs engineered in vitro from both primary cells and cell lines. Excitability is represented by rheobase (R(50), units = V/mm) and chronaxie (C(50), units = microseconds) values, with lower values for each indicating greater excitability. Adult skeletal muscle is the most excitable (R(50) ~ 0.29, C(50) ~ 100); chronically denervated whole muscles (R(50) ~ 2.54, C(50) ~ 690) and muscle engineered in vitro from cell lines (C2C12 + 10T1/2) (R(50) ~ 1.93, C(50) ~ 416) have exceptionally low excitability; muscle engineered in vitro from primary myocytes (R(50) ~ 0.99, C(50) ~ 496) has excitability similar to that of day 14 neonatal rat muscle (R(50) ~ 0.65, C(50) ~ 435); stimulated-denervated muscles retain excellent excitability when chronically electrically stimulated (R(50) ~ 0.40, C(50) ~ 100); and neonatal rat muscle excitability improves during the first 6 weeks of development, steadily approaching that of adult muscle.