Altered dopamine signaling in naturally occurring maternal neglect

Abstract

Background: Child neglect is the most common form of child maltreatment, yet the biological basis of maternal neglect is poorly understood and a rodent model is lacking.

Methodology/principal findings: The current study characterizes a population of mice (MaD1) which naturally exhibit maternal neglect (little or no care of offspring) at an average rate of 17% per generation. We identified a set of risk factors that can predict future neglect of offspring, including decreased self-grooming and elevated activity. At the time of neglect, neglectful mothers swam significantly more in a forced swim test relative to nurturing mothers. Cross-fostered offspring raised by neglectful mothers in turn exhibit increased expression of risk factors for maternal neglect and decreased maternal care as adults, suggestive of possible epigenetic contributions to neglect. Unexpectedly, offspring from neglectful mothers elicited maternal neglect from cross-fostered nurturing mothers, suggesting that factors regulating neglect are not solely within the mother. To identify a neurological pathway underlying maternal neglect, we examined brain activity in neglectful and nurturing mice. c-Fos expression was significantly elevated in neglectful relative to nurturing mothers in the CNS, particularly within dopamine associated areas, such as the zona incerta (ZI), ventral tegmental area (VTA), and nucleus accumbens. Phosphorylated tyrosine hydroxylase (a marker for dopamine production) was significantly elevated in ZI and higher in VTA (although not significantly) in neglectful mice. Tyrosine hydroxylase levels were unaltered, suggesting a dysregulation of dopamine activity rather than cell number. Phosphorylation of DARPP-32, a marker for dopamine D1-like receptor activation, was elevated within nucleus accumbens and caudate-putamen in neglectful versus nurturing dams.

Conclusions/significance: These findings suggest that atypical dopamine activity within the maternal brain, especially within regions involved in reward, is involved in naturally occurring neglect and that MaD1 mice are a useful model for understanding the basis of naturally occurring neglect.

Figure 1. Maternal neglect rate in MaD1 mice.

Maternal neglect rate (neglect leads to death of all pups in a litter) was significantly higher in MaD1 relative to Outbred-S mice when examined over 12 generations (A). Birth rate (number of male-female pairings relative to number of litters born) is almost identical between MaD1 and Outbred-S mice when examined over 12 generations (B). When raising a second litter (examined in Generation 6), previously neglectful mice exhibited significantly higher levels of maternal neglect relative to previously nurturing MaD1 mice (C). Bars represent means±SE. ***  = p<0.001.

Figure 2. Possible risk factors for maternal neglect in MaD1 mice across multiple generations.

When examined for the first litter in on postpartum Day 0, neglectful MaD1 mice weighed significantly less relative to their nurturing sisters (A) and average pup weight was significantly lower for pups born to neglectful mothers (B). Litter size was smaller for neglectful mice, but levels did not reach significance(C). Prior to mating, decreases in self-grooming (D) and increases in activity (E) were seen in neglectful relative to nurturing mice (D). For A–C, data from Generations (G) 12 and 15 were combined and for D and E, data from G6, G12, and G15 were combined. Time in light in a light/dark box test, did not differ between mice that would become either neglectful or nurturing when examined in G15 (F). Mice were first examined for anxiety at age 50 when they were group housed and then again after being singly housed for one week. In G17, neglectful mice spent significantly less time floating in the forced swim test relative to nurturing mice (G). Bars represent means±SE. *  = p<0.05; ***  = p<0.001.

Figure 3. Pups born to previously neglectful mice elicit decreases in maternal care.

When raised by either previously nurturing MaD1 mice or Outbred-S mice, pups that were born to previously neglectful MaD1 mothers received significantly higher levels of maternal neglect relative to pups born to previously nurturing MaD1 mice (A). Similarly, the proportion of pups weaned was significantly lower for pups born to previously neglectful MaD1 mothers when raised by either previously nurturing MaD1 mothers or Outbred-S mothers (B). Previously neglectful (relative to nurturing) MaD1 mice weaned a lower proportion of pups when the pups were from nurturing mothers, but the differences did not reach significance (p = 0.071). Bars represent means±SE. *  = p<0.05; **  = p<0.01.

Figure 4. Cross-fostered offspring raised by previously neglectful (relative to nurturing) MaD1 mothers show deficits as adults.

When results from all offspring were combined, previously neglectful MaD1 mothers negatively impacted offspring adult performance in terms of body weight pre-mating (A), flipping rate pre-mating (B), body weight on postpartum Day 0 (C), and number of maternal defense attacks (D). Additionally, when just offspring from previously nurturing MaD1 mothers were examined, they exhibited significantly lower levels self-grooming pre-mating when raised by a neglectful mother (E). When just offspring from Outbred-S mothers were examined as adults, deficits in terms of proportion of pups that survive to postpartum Day 10 (a marker of maternal neglect) (F), and the average weight of pups on postpartum Day 10 (G) were observed when these mice were raised by neglectful MaD1 mothers. Bars represent means±SE. *  = p<0.05; **  = p<0.01.

Figure 5. Altered c-Fos expression with maternal neglect.

Heightened c-Fos expression in dopamine releasing and responding regions in neglectful (N = 8) relative to nurturing (N = 9) mice. Significantly higher levels of c-Fos are found in ZI, VTA, and substantia nigra (SN), all of which are involved in dopamine production. Both nucleus accumbens shell (AcS) and core (AcC), which respond to dopamine signaling, also show increased c-Fos in neglectful mice. Other regions examined for c-Fos are shown in Table 1. Bars represent means±SE. *  = p<0.05; **  = p<0.01.

Figure 6. Altered pTH expression with maternal neglect.

Example of heightened pTH immunoreactivity in ZI in neglectful relative to nurturing mice is shown in (A). Significant elevations of pTH-ir area are found in ZI, but not other dopamine producing regions, in neglectful mice (N = 7) in comparison to nurturing dams (8) (B). TH-ir area does not differ between neglectful (N = 8) and nurturing (N = 8) mice. ZI = zona incerta, VTA = ventral tegmental area, A14 POA = A14 region of preoptic area. Bars represent means±SE. *  = p<0.05.

Figure 7. Altered pDARPP-32 expression with maternal neglect.

Previously neglectful dams (N = 5) have significantly darker optical density of pDARPP-32 expression in Ac and CP when compared to previously nurturing dams (N = 6). Ac = nucleus accumbens, LS = lateral septum, CP = caudate-putamen, BST = bed nucleus of stria terminalis, dorsal, CeA = central amygdala. Bars represent means±SE. *  = p<0.05.