Carrot production is constrained by high levels of heat stress during the germination stage in many global regions. Few studies have been published evaluating the effect of heat stress on carrot seed germination or screening for genetic heat stress tolerance. The objectives of this study were to evaluate the response of diverse carrot germplasm to heat stress, identify heat-tolerant germplasm that may be used by plant breeders, and define the appropriate temperature for assessing heat tolerance in germinating carrot seed. To identify an appropriate screening temperature, three commercial hybrids and an open pollinated variety were evaluated at five temperatures (24, 32.5, 35, 37.5, and 40 °C). In preliminary studies, 35 °C was identified as the optimal temperature for screening heat tolerance of carrot seed. Cultivated and wild carrot plant introductions (PIs) (n = 270) from the U.S. Department of Agriculture (USDA) National Plant Germplasm System (NPGS) representing 41 countries, inbred lines from the USDA Agricultural Research Service (n = 15), and widely grown commercial hybrids (n = 8) were evaluated for heat tolerance under heat stress and nonstress conditions (35 °C and 24 °C, respectively) by calculating absolute decrease in percent germination (AD), inhibition index (II), relative heat tolerance (RHT), and heat tolerance index (HTI). All measurements of heat tolerance identified significant differences among accessions; AD ranged from −13.0% to 86.7%, II ranged from 35.7% to 100.0%, RHT ranged from 0 to 1.36, and HTI ranged from 0.0 to 1.45. The broad-sense heritability (H2) calculations ranged from 0.64 to 0.86 for different traits, indicating a moderately strong genetic contribution to the phenotypic variation. Several wild carrot accessions and inbred lines displayed low levels of heat tolerance, whereas cultivated accessions PI 643114 (United States), PI 652400 and PI 652403 (Turkey), PI 652208 (China), and PI 652403 (Russia) were most heat tolerant. This is the first evaluation of heritability for heat stress tolerance during carrot seed germination, the first measure of HTI, and the first correlation calculation between heat and salt tolerance during germination in carrot.