Prefrontal cortex is associated with the rapid onset of parental behavior in inexperienced adult mice (C57BL/6)

Highlights

• 15 min of exposure to pups increases c-Fos in the brain of parental mice.

• The prelimbic cortex is associated with the immediate onset of parental behavior.

• High c-Fos in accumbens and amygdala was associated with the processing of pups.

• c-Fos expression in the MPOA was not associated with mouse parental behavior

Abstract

There is significant variability in the immediate behavioral response displayed by inexperienced adult mice when exposed to pups for the first time. The aim of this study was to determine which brain regions were engaged (higher c-Fos-immunoreactivity, c-Fos-ir) when virgin females, that were exposed to pups for 15 or 60 min, displayed full parental behavior (FPB), partial parental behavior (PPB), or non-parental behavior (NPB), or virgin males displayed PPB or infanticidal behavior (IB). The number of c-Fos-ir neurons in the prelimbic cortex (PL) was higher in parental females than in the NPB group (after a 15-min exposure), and the group not exposed to pups (NE). C-Fos expression in the nucleus accumbens (NA) was increased in most groups of females exposed to pups compared to NE. Higher c-Fos-ir was also found in the shell subregion of the NA in infanticidal males, compared to males NE. The cortical (CoA) and medial (MA) amygdala also showed higher c-Fos-ir in parental females compared to NE animals. However, PPB and IB male groups also exhibited higher c-Fos-ir in the CoA and MA compared to the NE group. The expression of c-Fos in the different subregions of medial preoptic area and the ventromedial nucleus of the hypothalamus was not specifically associated with either parental or infanticidal behavior. No brain activation in males was specifically associated with infanticidal behavior. Our results suggest that 15 min of exposure to pups is enough to detect brain regions associated with parental behavior (PL) or pups processing (NA, MA, CoA) in mice. The PL might participate in the immediate onset of parental behavior in virgin females, coordinating and planning its rapid execution.

Introduction

Parental behavior in rodents involves several behavioral components such as pup retrieval, crouching postures, nest building and licking/grooming [38,51,52], which contribute to the survival of the offspring [41,45,46]. Although mothers are commonly responsible for taking care of the young, both males and females express parental behavior in different social contexts and hormonal/physiological states. In some species, pup-naive virgin females and males can display parental behavior in non-reproductive contexts [39,51,52]. The induction of parental behavior by continuous exposure to pups is commonly called pup-induced parental behavior or sensitization [40,52]. However, there are inter and intraspecific differences in the latency to display parental behavior among the different rodent species [1,19,33,47,52]. For example, while most CB57BL/6 female mice can display parental behavior almost immediately or sensitize rapidly after 30 min to 3 h of exposure to pups, males from most mouse strains are generally infanticidal or non-parental (ignore or neglect the pups) [1,16,17,25,31,38,43].

Thus, the immediate behavioral response toward pups is extremely variable in mice, and can be either completely parental (all behavioral components of parental behavior are displayed), partially parental (only some of the behavioral components of parental behavior are displayed), non-parental (only one or none of the behavioral components of parental behavior are displayed) or infanticidal (animals attack or hurt pups). Therefore, mice provide a good opportunity to discriminate brain regions that might be specifically engaged in parental or infanticidal behavior from those engaged in just the processing of pup-related stimuli and other social or non-social stimuli. Previous studies [7,58] investigated the expression of c-Fos only in female mice that showed full parental behavior and after 30 min of exposure to pups. Some of these studies also did not find behavioral variability in the immediate behavioral response to pups [7], something that differed from other studies, including ours [1,25,33]. Besides, all those previous studies [7,24,[57], [58], [59]] compared c-Fos expression between groups of parental/infanticidal mice and groups of mice not exposed to pups, or in different reproductive contexts. A control group of non-parental virgin mice exposed to pups was never used. Therefore, the expression of c-Fos found in those studies might not be specifically associated with parental or infanticidal behavior, but just with the exposure to pups or the presence of a different physiological state. Investigating the specificity of previous findings is one of the goals of the current study.

Mice also offer an opportunity to explore rapid neural changes that occur when animals are responding differently toward the same stimulus. The process of rapid sensitization gives us also the opportunity to compare brain regions engaged in the initial stages of the interaction with pups (first 15 min) from those engaged after longer interaction with them (60 min or several days). The immediate early gene c-fos is rapidly and transiently expressed in neurons in response to a variety of extracellular stimuli. Transcriptional activation of the gene occurs within minutes of stimulation, with levels of the nuclear protein c-Fos increasing 30–40 min later [4,9,20,35,36,54]. However, the temporal changes and maintenance of c-Fos expression depend on the areas of the brain studied [11,36,42,60]. For example, c-Fos mRNA expression was maximal in the dentate gyrus, cortical regions, accumbens, amygdala, septum, and lateral habenula before or at 60 min of an acute stress [9], morphine and convulsant administration [36,60]. Besides, other authors, that used similar time of sacrifice and procedures to us, found increased c-Fos in lateral habenula and raphe, but not in the medial preoptic nucleus, at 70 min after the beginning of the exposure to pups [11]. The expression of the c-Fos mRNA decline after that to low or basal levels at 1.5 or 2 h post stimulus [9]. The protein c-Fos was also observed to be maximal around 1 h in several brain regions [10,11]. Thus, the time of sacrifice and perfusion after stimulus presentation can differently affect the observed c-Fos expression in the different brain regions [11,60]. In the current study we decided to sacrifice animals after 1 h of the presentation of the stimuli (pups, object, or pups inside object) to detect brain regions rapidly, and perhaps transiently, engaged in the onset of parental behavior. Besides, we investigated for the first time if 15 min of exposure to pups was enough to increase c-Fos-ir neurons in the brain.

We analyzed several brain regions that included the prelimbic (PL) and infralimbic (IL) cortex, the medial preoptic area (MPOA), and the nucleus accumbens (NA). The PL and IL cortex might be critical for the recognition of stimulus, attentional selection, decision making, behavioral flexibility and planning of the immediate parental or infanticidal response [2,49,50]. The MPOA is critical for the expression of parental behavior in parturient females as well in sensitized females and males of several species [7,10,23,24,26,27,42,58,59]. However, the MPOA has several subareas and some of them have been associated also with infanticidal behavior or the inhibition of parental behavior [24,53]. Moreover, the NA has been involved in the processing of information related with the offspring and the facilitation of parental behavior [8,18,32,22,28,29,41,44,47,48]. However, this area has also been classically associated with the processing of novelty and the mediation of motivation to approach or avoid appetitive and aversive stimuli [5,34]. In the current study we investigated which brain regions expressed c-Fos associated with the rapid decision to take care of (parental behavior) or attack (infanticidal behavior) the pups. We expected that non-parental, parental and infanticidal animals would show different pattern of expression of c-Fos in these brain regions and those differences would be evident rapidly.

We found that 15 min of exposure to pups was enough to detect brain regions associated with parental behavior or pup processing. Besides, the PL was the only brain region specifically associated with parental behavior suggesting it plays a role in the immediate onset of parental behavior, perhaps coordinating and planning its rapid execution. Other areas of the brain, commonly associated with the stimulation (MPOA, NA), or inhibition of parental behavior (i.e. MA, CoA, and ventromedial nucleus of the hypothalamus, VMH) [6,14,37,57] were not specifically associated with parental or infanticidal behavior.