Developmental programming of somatic growth, behavior and endocannabinoid metabolism by variation of early postnatal nutrition in a cross-fostering mouse model

<div><p>Background</p><p>Nutrient deprivation during early development has been associated with the predisposition to metabolic disorders in adulthood. Considering its interaction with metabolism, appetite and behavior, the endocannabinoid (eCB) system represents a promising target of developmental programming.</p><p>Methods</p><p>By cross-fostering and variation of litter size, early postnatal nutrition of CB6F1-hybrid mice was controlled during the lactation period (3, 6, or 10 pups/mother). After weaning and redistribution at P21, all pups received standard chow ad libitum. Gene expression analyses (liver, visceral fat, hypothalamus) were performed at P50, eCB concentrations were determined in liver and visceral fat. Locomotor activity and social behavior were analyzed by means of computer-assisted videotracking.</p><p>Results</p><p>Body growth was permanently altered, with differences for length, weight, body mass index and fat mass persisting beyond P100 (all 3>6>10,p<0.01). This was paralleled by differences in hepatic IGF-I expression (p<0.01). Distinct gene expression patterns for key enzymes of the eCB system were observed in fat (eCB-synthesis: 3>6>10 (DAGLα p<0.05; NAPE-PLD p = 0.05)) and liver (eCB-degradation: 3>6>10 (FAAH p<0.05; MGL p<0.01)). Concentrations of endocannabinoids AEA and 2-AG in liver and visceral fat were largely comparable, except for a borderline significance for higher AEA (liver, p = 0.049) in formerly overfed mice and, <i>vice versa</i>, tendencies (p<0.1) towards lower AEA (fat) and 2-AG (liver) in formerly underfed animals. In the arcuate nucleus, formerly underfed mice tended to express more eCB-receptor transcripts (CB1R p<0.05; CB2R p = 0.08) than their overfed fellows. Open-field social behavior testing revealed significant group differences, with formerly underfed mice turning out to be the most sociable animals (p<0.01). Locomotor activity did not differ.</p><p>Conclusion</p><p>Our data indicate a developmental plasticity of somatic growth, behavior and parameters of the eCB system, with long-lasting impact of early postnatal nutrition. Developmental programming of the eCB system in metabolically active tissues, as shown here for liver and fat, may play a role in the formation of the adult cardiometabolic risk profile following perinatal malnutrition in humans.</p></div

Concentrations of endocannabinoids anandamide (AEA) and 2-arachidonoyl glycerol (2 AG) as well as arachidonic acid (AA) in visceral (perigonadal) adipose tissue.

<p>Except for tendencies towards lower AEA concentrations in formerly underfed animals (P50: 10 vs. 3/6 p = 0.089; P100: 10 vs.3/6 p = 0.063) no differences between former nutrition groups were detected at P50 and P100, resp. n = 4–6 males per nutrition group.</p

Relative gene expression of eCB-metabolizing enzymes in visceral (perigonadal) adipose tissue at P50 in relation to early postnatal nutrition.

<p>n = 4–6 males per nutrition group, * p<0.05 (ANOVA).</p

BMI (Fig. 2A) and perigonadal fat mass (Fig. 2B) at P50 and P100 in relation to postnatal nutrition and gender.

<p>Group differences for both parameters tended to further increase with age. If not indicated otherwise, displayed significance levels (* p<0.05, ** p<0.01, *** p<0.001) refer to comparisons between all three nutrition groups (ANOVA). P50: n = 4–10 animals per nutrition group and gender; P100: n = 5–10 animals per nutrition group and gender.</p

Relative gene expression of eCB receptors in the hypothalamic arcuate nucleus at P50 in relation to early postnatal nutrition.

<p>n = 4–5 males per nutrition group, * p<0.05 (ANOVA).</p

Relative gene expression of eCB-metabolizing enzymes in the liver at P50 in relation to early postnatal nutrition.

<p>If not indicated otherwise, displayed significance levels (* p<0.05, ** p<0.01) refer to comparisons between all three nutrition groups (ANOVA). n = 4–6 males per nutrition group.</p

Relative gene expression of IGF-I in the liver at P50 in relation to early postnatal nutrition.

<p>If not indicated otherwise, displayed significance levels (** p<0.01, *** p<0.001) refer to comparisons between all three nutrition groups (ANOVA). n = 4–6 males per nutrition group.</p

Body weight in grams (y-axis) and age in days of postnatal development (x-axis), longitudinal data of a P100 cohort.

<p>Note that differences between the nutrition groups (3, 6, and 10 pups per mother, respectively) persisted after weaning and redistribution at P21 (dotted line), after which mice had free access to standard food.</p

Concentrations of endocannabinoids anandamide (AEA) and 2-arachidonoyl glycerol (2 AG) as well as arachidonic acid (AA) in liver tissue.

<p>In addition to higher AEA (P100, p = 0.049) in formerly overfed animals and a trend towards lower 2-AG concentrations in formerly underfed animals (P50, p = 0.072), we observed a significantly reduced total amount of the eCB component arachidonic acid (AA) in the former overfed group (3 vs. 6/10 p = 0.023). n = 4–7 males per nutrition group.</p

Open field experiment investigating social behavior at P45-P50.

<p>During a 10 min recording time, substantial differences were observed for parameters assessing social interaction with an unknown partner animal. Although formerly overfed males exhibited the highest partner visiting frequency, they spent the least time in the vicinity of the partner. Locomotor activity was not different between the nutrition groups. Data are presented as mean ±SEM (n = 6–8 males per nutrition group). If not indicated otherwise, displayed significance levels (* p<0.05, ** p<0.01, *** p<0.001) refer to comparisons between all three nutrition groups (ANOVA).</p