Left ventricular diastolic filling is a fundamental constituent of cardiac performance. Diastolic function in both adults and children can be routinely assessed by radionuclide ventriculography (RNV). It has previously been shown that factors such as heart rate (HR) and age can significantly modify diastolic performance in adults, thus limiting the clinical applicability of RNV diastolic indices. The aim of this study was to investigate various factors that may affect diastolic function in childhood. Seventy-nine children, aged 40 days to 15 years, were enrolled in the study; their HR ranged from 45 to 160 beats per minute (bpm). All had intact cardiac function and were submitted to baseline RNV prior to chemotherapy initiation for malignancies. Using stepwise linear regression analysis, HR was identified as the major factor affecting RNV diastolic indices during growth. Applying univariate regression models, diastolic indices were corrected for a referrence HR of 100 bpm; this substantially reduced variability of RNV diastolic indices along age increments, allowing for the establishment of reference ranges. In conclusion, HR was shown to be the major determinant of RNV diastolic indices during growth. Adjustment for this variable alone can offer reference ranges for the assessment of left ventricular filling in childhood.