Previous research has demonstrated the fundamental role of the caregiver in shaping toddlers’ pain-related distress responses. One caregiver factor that has remained largely understudied in the context of toddler acute pain is cardiac responses during their children’s medical procedures. The aim of the current study was to examine the bidirectional associations between caregivers’ respiratory sinus arrhythmia (RSA) and toddlers’ behavioural pain-related distress during a vaccination appointment. Toddler behavioural distress immediately before the needle negatively predicted caregiver RSA immediately after the needle. Caregiver RSA at 1-minute post-needle positively predicted toddler behavioural distress scores at 2-minutes post-needle. The current results underscore the importance of optimal parent-led pain management strategies being implemented during routine pediatric vaccinations.