Safety has been defined as the absence of threat and stimuli never associated with an aversive event (unconditioned stimulus, US) can inhibit conditioned defensive responses. Relief is a positive response elicited by the termination of an aversive US and stimuli presented upon the moment of relief elicit appetitive conditioned responses. Unclear remains whether the threat absence and threat termination share inhibitory mechanisms or rather these two types of safety are distinct. Fifty-eight participants learned that one stimulus (forward CS+) was shortly presented before a mildly painful electric stimulation (US), one stimulus (backward CS+) was presented shortly after the US, and one stimulus (CS-) was never associated with the US. During a summation test, forward CS+ was presented in compound with either the backward CS+ or the CS-. Conditioned defensive responses were successfully acquired on both verbal and physiological responses meaning that forward CS+ compared to both CS- and backward CS+ was rated more aversive and elicited stronger physiological responses. During summation test, conditioned physiological defensive responses were significantly and comparably attenuated by both backward CS+ and CS-, but inhibition of startle potentiation by the relief-associated stimulus was not evident during the early test trials. In summary, conditioned defensive responses can be inhibited by signals of threat absence (CS-) and threat termination (backward CS+). However, the underlying mechanisms of these two signals may differ.