Toll-like receptors (TLR) are innate immune receptors typically activated by microbial-associated molecular patterns (MAMPs) during infection or damage-associated molecular patterns (DAMPs) as a result of tissue injury. Recent findings suggest that TLR2 and TLR4 signaling play important roles in developmental and adult neuroplasticity, and in learning and memory. In addition, activation of TLR2 and TLR4 worsens ischemic injury to the heart and brain in animal models of myocardial infarction and stroke. TLR activation is also implicated in thermoregulation and fever in response to infection. However, it is not known whether TLRs participate in the regulation of the sympathetic and/or parasympathetic components of the autonomic nervous system (ANS). Here we provide evidence that TLR2 and TLR4 influence autonomic regulation of heart rate (HR) body temperature and energy metabolism in mice. We show that mice lacking TLR2 or TLR4 exhibit reduced basal HR, which results from an increase of parasympathetic tone. In addition, thermoregulatory responses to stress are altered in TLR2-/- and TLR4-/- mice, and brown fat-dependent thermoregulation is altered in TLR4-/- mice. Moreover, TLR2-/- and TLR4-/- mice consume less food and exhibit a greater mass compared to wild type mice. Collectively, our findings suggest important roles for TLR2 and TLR4 in the ANS regulation of cardiovascular function, thermoregulation, and energy metabolism.