Ectothermic animals are susceptible to temperature changes such as cold shock with seasons. To survive through a cold shock or season, ectotherms have developed unique strategies. Our interest is focusing on the modulation of physiological functions during cold shock and prolonged cold exposure in the fruit fly. We use Drosophila melanogaster as a model system to investigate cardiac function in response to modulators (5-HT—serotonin, Ach—acetylcholine, OA—octopamine, DA—dopamine and a cocktail of modulators) in acute cold shock and chronic cold shock conditions. Semi-intact larvae are used to provide direct access to the modulators of known concentration in a defined saline. The results show that 10 µM 5HT is the only modulator which maintains heart rate for larva raised at 21 °C and then exposed to acute cold shock (10 °C). The modulators 1 µM OA, 10 µM 5HT, 1 mM Ach, 10 µM Ach and a cocktail of modulators (at 10 µM) increased the heart rate significantly in larvae which were cold conditioned (10 °C for 10 days). HPLC analysis indicated both OA and 5-HT decreased in chronic cold conditioning. The larvae maintain heart function in the cold which may be contributed by low circulating levels of modulators. The larval heart responds better to 5-HT, OA, and Ach in conditioned cold than for acute cold, suggesting some acclimation to cold.