Signs Righting reﬂex Negative geotaxis Cliﬀ avoidance Acoustic startle Supported standing Unsupported standing PND 1 PND 2 PND 3 PND 4 PND 4 PND 5 PND 6 PND 6 PND 7 PND 9 PND 10 PND 11 PND 7 PND 8 PND 9 PND 10 PND 15 PND 16 PND 17 PND 18 Control 0 28.8 53.6 96.4 23 56 82 10.7 50 0 5.4 53.6 0 35.7 62.5 73.2 0 33.9 74.9 100 EVA 10.7 80.4 100 100 25 54 85 30.3 70 16.1 42.9 100 26.8 7.8 100 100 44.6 100 100 100 Control 0 39.3 76.6 94.6 35 70.2 100 14.3 55 0 14.3 41 0 36 59 89.3 0 39.2 66.1 89.3 EVA 5.4 71.4 100 100 38 71 100 26.7 73.2 19.6 53.6 93 26.8 73.2 100 100 21.4 75 100 100 4 Evidence-Based Complementary and Alternative Medicine Table 1: Eﬀects of maternal dietary EVA supplementation on the onset and completion of neuromotor skill and neurological reﬂex acquisi- tion in rat pups. Day of onset1 Day of completion1 Signs2 Male Female Male Female Control EVA Control EVA Control EVA Control EVA Righting reﬂex 2.0 (0.01) 1.0 (0.05) 2.0 (0.02) 1.1 (0.07) 4.9 (0.25) 3.1 (0.12) 4.9 (0.26) 3.1 (0.12) Negative geotaxis 4.1 (0.18) 4.0 (0.20) 4.0 (0.20) 4.0 (0.20) 6.9 (0.27) 6.8 (0.28) 6.5 (0.30) 6.5 (0.30) Cliﬀ avoidance 6.6 (0.30) 6.3 (0.26) 6.9 (0.20) 6.9 (0.33) 8.0 (0.35) 6.6 (0.28) 7.9 (0.32) 7.9 (0.35) Acoustic startle 11.0 (0.45) 9.4 (0.40) 11.0 (0.45) 9.4 (0.40) 12.0 (0.50) 11.0 (0.45) 12.0 (0.50) 11.6 (0.51) Supported standing 8.0 (0.34) 7.6 (0.33) 8.0 (0.34) 7.6 (0.33) 11.2 (0.65) 10.0 (0.00)∗ 10.9 (0.48) 9.1 (0.34)∗ Unsupported standing 15.9 (0.66) 14.9 (0.65)∗ 15.8 (0.56) 15.0 (0.70) 17.9 (0.66) 15.8 (0.30)∗ 17.9 (0.66) 16.1 (0.40)∗ 1 Data are expressed as mean ± standard error of man (SEM). 2The acquisition of various reﬂexes (yes/no) was assessed at speciﬁc postnatal ages, for example, Righting Reﬂex at PND 1–5, Negative Geotaxis at PND 4–6, Cliﬀ Avoidance at PND 6–8, Acoustic Startle at PND 9–12, Supported standing at PND 7–11, and Unsupported standing at PND 15–18. ∗Signiﬁcant (P ≤ 0.05) diﬀerences between control and EVA groups (within gender) in the onset or completion of physical and neurological signs of development. Table 2: Percentage of rat pups possessing an acquired neuromotor and neurological reﬂex on speciﬁc postnatal days. % of pups Male Female Diet eﬀect (P value) Gender eﬀect (P value) NS: not signiﬁcant. PND: postnatal day, recorded as the ﬁrst PND where a signiﬁcant diﬀerence (P ≤ 0.05) was noted. of pups showing these reﬂex activities over time was not diﬀerent (data not shown). 4. Discussion The primary aim of our study was to investigate the eﬀects of long-term maternal dietary EVA supplementation in the diet of female rats prior to mating, and for the duration of pregnancy and lactation, on the physical, reﬂex, and neuro- motor development of the progeny. We employed a battery of behavioral tests that reﬂect the developmental maturation of the central nervous system [22] and assessed various parameters of physical development. We used long-term maternal and paternal supplementation with EVA as several studies identify the importance of diet on fetal and postnatal development through diet-mediated epigenetic regulation 0.002 <0.001 <0.001 0.024 0.019 0.035 0.004 0.003 0.004 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001 0.013 NS NS NS NS NS NS NS NS NS NS NS NS NS NS NS 0.039 0.019 NS NS 0.013