Background and Aims: Nocturnal hypoglycemia is most feared in children with diabetes, though often unrecognized. Heart rate variability (HRV) reflects activation of the autonomic nervous system. As hypoglycemia and declining glucose induce sympathoadrenal activation, changes of HRV could be assumed. Hence, the aim of this study was to evaluate, if HRV changes occur before the onset of nocturnal hypoglycemia in children with type 1 diabetes (T1D). Methods: Children aged 1 to 18 y, with T1D for >6 months, participated in an observational study using continuous glucose monitoring (CGM) during 5 days. Simultaneously, Holter ECG was recorded each night. HRV parameters such as time (SDNN) and frequency (VLF and total power) domain and sample entropy were calculated using a 6min moving window from 90mins before until the onset ( = 0mins) of nocturnal hypoglycemia (sensor glucose level <3.9mmol/l for ≥15mins). Mean HRV parameters at several timepoints were compared to timepoint 0 using a Kruskal‐Wallis test. Results: Twenty‐five children (11f, mean (SD) age 13.5y (2.5) participated in the study. 33 hypoglycemic events with concomitant ECG recording occurred. Mean SDNN increased from 79ms at ‐70mins and 83ms at ‐30mins to 89ms at 0mins (p = 0.002 and 0.01), as did VLF (from 3158ms and 4366ms to 5153ms; p = 0.0001, 0.0002) and total power (from 7960ms and 9663ms to 10700ms; p = 0.0002, 0.0081). Sample entropy decreased from 1.76 and 1.75 to 1.65 (p = 0.001, 0.001). Conclusions: Significant changes of multiple HRV variables as early as 70 mins before the occurence of nocturnal hypoglycemia were documented. This indicates that HRV changes may be used to help predict nocturnal hypoglycemia.