Fear response of rat pups to a non-aversive social stimulus: Evidence for the involvement of memory processes.

Learning processes in rats during early development are importantly mediated by the mother, which represents the primary source of environmental information. This study aimed to determine whether aversive early experiences can induce the expression of pups' fear responses toward a non-aversive stimulus as a consequence of a memory process. First, we determined pups' fear responses toward an anesthetized female after being exposed to this stimulus or an empty cage together with their mothers from Postnatal Day (PNDs) 1 to 4. Second, we evaluated if the administration of the protein synthesis inhibitor cycloheximide (CHX; 0.2 mg/kg, subcutaneously (sc).) disrupted the reconsolidation processes and abolished the fear response on PND 9. Only female pups previously exposed to the female intruder expressed fear responses toward an anesthetized female on PND 8. CHX administration to female pups immediately after exposure to an anesthetized female on PND 8 suppressed fear responses on PND 9, indicating that the fear expression was the result of a memory process, probably mediated by the mother. These findings demonstrated that early experiences can shape responses to social stimuli in a sex-dependent manner and emphasize the critical role of the mother in influencing fear learning in a social context.

Raising overlapping litters: Differential activation of rat maternal neural circuitry after interacting with newborn or juvenile pups.

The maternal behaviour of a rat dynamically changes during the postpartum period, adjusting to the characteristics and physiological needs of the pups. This adaptation has been attributed to functional modifications in the maternal circuitry. Maternal behaviour can also flexibly adapt according to different litter compositions. Thus, mothers with two overlapping litters can concurrently take care of neonate and juvenile pups, mostly directing their attention to the newborns. We hypothesised that the maternal circuitry of these mothers would show a differential activation pattern after interacting with pups depending on the developmental stage of their offspring. Thus, we evaluated the activation of several areas of the maternal circuitry in mothers of overlapping litters, using c-Fos immunoreactivity as a marker of neuronal activation, after interacting with newborns or juveniles. The results showed that mothers with overlapping litters display different behavioural responses towards their newborn and their juvenile pups. Interestingly, these behavioural displays co-occurred with specific patterns of activation of the maternal neural circuitry. Thus, a similar expression of c-Fos was observed in some key brain areas of mothers that interacted with newborns or juveniles, such as the medial preoptic area and the nucleus accumbens, whereas a differential activation was quantified in the ventral region of the bed nucleus of the stria terminalis, the infralimbic and prelimbic subregions of the medial prefrontal cortex and the basolateral and medial nuclei of the amygdala. We posit that the specific profile of activation of the neural circuitry controlling maternal behaviour in mothers with overlapping litters enables dams to respond adequately to the newborn and the juvenile pups.

The Expression of Hypoxia-Induced Gene 1 (Higd1a) in the Central Nervous System of Male and Female Rats Differs According to Age.

HIGD1A (hypoxia-induced gene domain protein-1a), a mitochondrial inner membrane protein present in various cell types, has been mainly associated with anti-apoptotic processes in response to stressors. Our previous findings have shown that Higd1a mRNA is widely expressed across the central nervous system (CNS), exhibiting an increasing expression in the spinal cord from postnatal day 1 (P1) to 15 (P15) and changes in the distribution pattern from P1 to P90. During the first weeks of postnatal life, the great plasticity of the CNS is accompanied by cell death/survival decisions. So we first describe HIGD1A expression throughout the brain during early postnatal life in female and male pups. Secondly, based on the fact that in some areas this process is influenced by the sex of individuals, we explore HIGD1A expression in the sexual dimorphic nucleus (SDN) of the medial preoptic area, a region that is several folds larger in male than in female rats, partly due to sex differences in the process of apoptosis during this period. Immunohistochemical analysis revealed that HIGD1A is widely but unevenly expressed throughout the brain. Quantitative Western blot analysis of the parietal cortex, diencephalon, and spinal cord from both sexes at P1, P5, P8, and P15 showed that the expression of this protein is predominantly high and changes with age but not sex. Similarly, in the sexual dimorphic nucleus, the expression of HIGD1A varied according to age, but we were not able to detect significant differences in its expression according to sex. Altogether, these results suggest that HIGD1A protein is expressed in several areas of the central nervous system following a pattern that quantitatively changes with age but does not seem to change according to sex.

Incentive value of newborn pups relative to juveniles for mother rats raising overlapping litters.

In rats, successful mating during the postpartum estrus results in the temporal overlapping of successive litters within the maternal nest. Mothers with two overlapping-litters (OLM) simultaneously take care of neonate and juvenile pups; however, they mostly direct their attention to the neonates. We hypothesized that these differences reflect an adaptation to the specific characteristics and needs of the two litters and not a lack of interest in the juveniles. To test this hypothesis, we assessed the relative incentive value of newborns and juveniles for OLM in a preference test and compared it with that exhibited by mothers in early (EPM) and late (LPM) postpartum, which were raising only newborns or only juveniles, respectively. Results showed that OLM spent similar time in the newborns and juveniles compartments and did not prefer the newborns as did the EPM, however, similarly to them, OLM made more attempts to get access to the newborns than the juveniles. On the other hand, OLM and LPM did not exhibit a clear preference between the stimuli. These results indicate that both neonates and juveniles have incentive value for OLM, although these mothers invest more effort in the newborns. These results point out to a unique behavioral profile of OLM, which shows similarities with EPM and LPM on different behavioral measures. They also support the idea that motivational processes underlying maternal behavior are complex and dynamic, adapting the response of the mother to pups' needs and the context.

Dopaminergic activity mediates pups' over male preference of postpartum estrous rats.

Pups have greater incentive value than males for rats during the postpartum estrus (PPE); a period when females are both maternally and sexually motivated. Mesolimbic dopaminergic system has been proposed as a general motivational circuit; however in the literature it has been more related to the control of the motivational aspects of maternal than sexual motivation of females. Therefore, we aimed to assess the effect of antagonizing dopaminergic neurotransmission of PPE females on their preference for pups over a male. To achieve this objective we tested PPE rats in a Y-maze with three-choice chambers (one containing eight pups, the other a male and the last one no stimulus) after the systemic administration of the dopaminergic antagonist haloperidol (0.0; 0.025 or 0.05 mg/kg). Furthermore, to determine if this dopaminergic antagonist differentially affects maternal and sexual motivations when pups and male are not competing, we evaluated the effect of haloperidol in the preference of females for pups vs. a non-receptive female and for a male vs. a non-receptive female. In the preference test for pups vs. male, both doses of haloperidol decreased the time that females spent in pups' chamber while increased the time that they spent in male's chamber, resulting in a lack of preference between both incentives. Besides, haloperidol reduced the effort -attempts to get access to the stimuli- made by the females to obtain the pups. Conversely, 0.05 mg/kg of haloperidol did not affect the preference for both incentives when they were confronted to a non-receptive female. Together, these results indicate that the dopaminergic activity mediates pups' preference over male during the PPE and point toward a more relevant role of this system in females' behavioral output when incentives are competing.

Previous and recent maternal experiences modulate pups' incentive value relative to a male without affecting maternal behavior in postpartum estrous rats.

This study extends the behavioral analysis of the postpartum estrus (PPE) which represents a unique period in the female rat's lifetime when maternal and sexual motivations co-exist. The aim of this study was to explore how previous and recent maternal experiences influence the maternal responses to pups when confronted with a male in a preference test or when they are presented independently in the home cage. To achieve this objective, we firstly compared the maternal behavior in the home cage and the preference for pups or a male in a Y-maze of primiparous and multiparous females approximately twelve hours after delivery. No differences were observed in the active and passive components of the maternal behavior of primiparous and multiparous rats; however second-time mothers made more efforts to gain access to the pups and tended to spend more time with them in the Y-maze than maternally inexperienced dams. In a second experiment, we assessed the influence of recent maternal experience with pups on PPE females' behavior by comparing pups vs. male preference and maternal behavior of females that had experienced continuous or limited (approximately two hours) interaction with their litters after parturition was completed. PPE rats subjected to reduced interaction with their pups preferred the male, while females continuously exposed to pups chose them over the male. This change in females' preference was not accompanied by significant alterations of maternal performance in the home cage, although anogenital licking tended to decrease in females with limited mother-litter interaction. Together, the results of these experiments indicate that previous and recent maternal experiences influence the motivational responses of PPE females, and that these effects are more evident when both motivations compete.

Maternal and affective behaviors of lactating rats reared in overlapping litters.

Postpartum mating in rats gives rise to complex family units consisting of the mother and two overlapping litters. As a consequence, newborn pups of the second litter, since the moment they are born, acquire experience not only from interaction with the mother and age-matched littermates but also from interaction with older siblings. Newborn pups reared in overlapping litters (OLs) receive a different pattern of maternal stimulation compared to those reared in single litters (SL: one litter of same aged pups), as the mothers reduce some maternal behavior components and juvenile pups from the first litter develop maternal behavior. Since there is strong evidence showing that variations in maternal behavior are transmitted throughout generations, we hypothesized that the altered pattern of maternal stimulation received by OL reared females would modify their behavior during motherhood. To test this hypothesis maternal behavior, maternal aggression and experimental anxiety of dams reared under OL and SL conditions during the first postpartum week were compared. No differences were found between the groups in their maternal behavior and aggression. This result may be explained by the maternal behavior of the juveniles that could compensate for the deficits in the caregiving behaviors received by OL litters. However, a subtle temporal reorganization of the licking behavior was found in OL reared mothers, together with an increased anxiety-related behavior in the plus maze test. These results suggest dissociation in the effects provoked by early environmental alterations on different behavioral systems, and more importantly, that independently of their early family composition, both groups can cope effectively with the changing demands of the pups.

Different chemical fractions of fetal fluids account for their attractiveness at parturition and their repulsiveness during late-gestation in the ewe.

We have investigated whether the chemical components of fetal fluids (FFs), which elicit repulsion in late gestating ewes, are also those responsible for the attractiveness of fetal fluids at parturition. An aqueous fraction of FFs (A1), obtained after extraction with hexane, was tested for repulsion in late-pregnant ewes and for attraction at parturition. We also investigated if the repulsive and attractive characteristics of this A1 fraction were maintained after an additional extraction with dichloromethane (DCM, CH(2)Cl(2)) that produced two more fractions (aqueous/high polarity: A2 and dichloromethane/medium polarity: DCM). Thus, late-pregnant ewes were tested for repulsion of aqueous extracts of FFs (A1, A2 and DCM fractions) in a two-choice test of food preference, whereas parturient ewes were tested for attraction toward these same fractions in a two-choice test of licking warm spongy cloths. The A1 fraction was repulsive to late-pregnant ewes and attractive to parturient females. In contrast, neither the A2 nor the DCM fractions were repulsive to late-pregnant ewes, whereas both fractions were attractive to parturient ones. The discordance between the repulsive and attractive properties of the A2 and DCM fractions suggests that the attractiveness of FFs for parturient ewes and its repulsiveness for females outside the peripartum period depend on mixtures of substances that are at least partially different. Some compounds with high and medium polarity in the A2 and DCM fractions would act synergistically to generate the repulsiveness of FFs, whereas both high and medium polarity compounds can evoke attraction independently of each other.

Compulsive-like behaviour according to the sex and the reproductive stage of female rats.

The aim of the present study was to explore putative differences in the responses assessed in an animal model of obsessive-compulsive disorder (OCD) according to the sex and the reproductive cycle of female rats. The model consists of the induction of perseveration (repetitive choices of the same arm in a T-maze) by 8-OH-DPAT (1.0mg/kg). Males and females (pooled in all stages of their oestrous cycle) persevered after 8-OH-DPAT administration and no differences were observed between groups. During the oestrous cycle, this 5-HT(1A) agonist induced perseveration in metoestrus, dioestrus and prooestrus and reduced levels of this behaviour in oestrus. 8-OH-DPAT provoked perseveration in mid-gestation, an effect that was reduced in late-gestation and blocked during lactation. Reproductive cycle changes in the induced perseveration are discussed from the standpoint of the ovarian steroids' action on the serotoninergic system and on the bases of the variations in stress responsiveness along the reproductive cycle of the female. Present results validate the use of females in this model of OCD and could be relevant for studying the role of reproductive hormones in the pathophysiology of this disorder.