Joan I. Morrell

Comparison of two positive reinforcing stimuli: pups and cocaine throughout the postpartum period.

This set of experiments investigated the appetitive or motivational processes underlying the performance of maternal behavior. The place preference paradigm was adapted to simultaneously investigate the reinforcing properties of cocaine and pups for maternal, lactating dams. These modifications allowed the authors to assess which stimulus, either a 10 mg/kg s.c. injection of cocaine or 3 pups, had the strongest reinforcing value. At Postpartum Days 10 and 16, the dams preferred the cocaine cue-associated chamber, whereas the dams tested at Postpartum Day 8 preferred the pup cue-associated chamber. Overall, the data revealed an interaction between the postpartum period at testing and the exhibited preference for cocaine or pups. Further testing will investigate the neural circuitry underlying the appetitive processes of each stimulus.

Distribution and steroid hormone regulation of aromatase mRNA expression in the forebrain of adult male and female rats: A cellular‐level analysis using in situ hybridization

Many of the effects of gonadal steroid hormones in the male brain are due to the actions of the testosterone metabolite estradiol, which is synthesized by the actions of the P450 enzyme aromatase. Aromatase activity is present in regions of the preoptic area, hypothalamus, and limbic system. Levels of aromatase activity in the brain are highly dependent on gonadal steroid hormones in many brain regions, but not all. We examined the distribution of aromatase mRNA in adult male and female rat brains as well as the regulation of the levels of aromatase mRNA in the brains of males by gonadal steroid hormones using in situ hybridization. This method was performed using a 35S‐labelled cRNA probe, transcribed in vitro from the rat ovarian aromatase cDNA. In the adult male, many heavily labelled cells were found in the encapsulated bed nucleus of the stria terminalis (BNST), the medial preoptic nucleus (MPN), the ventromedial nucleus (VMN), the medial amygdala (mAMY), and the cortical amygdala (CoAMY). The regional distribution of aromatase mRNA was similar in females, but females tended to have a lower number of aromatase mRNA expressing cells in each region compared to males. Aromatase mRNA levels in the BNST, MPN, VMN, and mAMY tended to be lower in castrated males than in intact males, whereas aromatase mRNA levels were unaltered by castration in the CoAMY. The degree of reduction in mean levels of aromatase mRNA following castration does not simply account for the large changes measured in activity following castration. Examination of the entire population of individual cells expressing aromatase mRNA in castrated males suggests that aromatase mRNA may be regulated by steroid hormones differentially in specific populations of neurons within regions where activity is known to decrease following castration.

Preference for cocaine- versus pup-associated cues differentially activates neurons expressing either fos or cocaine- and amphetamine-regulated transcript in lactating, maternal rodents

We studied the neuronal basis of the motivational response to two powerful but radically different rewards-cocaine and maternal nurturing of pups in the postpartum rat (dam) which is in a unique motivational state. We used a place preference method designed to offer a choice between cues associated with a natural reinforcer (pups) and those associated with a pharmacologic reinforcer (cocaine). Using c-Fos or cocaine- and amphetamine-regulated transcript (CART) immunocytochemistry, we identified the neuronal groups that are activated when the dams expressed a preference for either cues-associated with pups or cues-associated with cocaine. Dams that preferred the cocaine-associated cues had more c-Fos positive neurons in medial prefrontal cortex, nucleus accumbens, and basolateral nucleus of amygdala than pup-associated cue preferring dams or control. Except for the accumbens, there was activation of neurons in these same regions with the pup-associated cue preference. In the nucleus accumbens only CART-immunoreactive (not c-Fos) neurons were activated with pup-cue preference. Notably, the medial preoptic area was the single area where greater activation of neurons was seen with a preference for pup-associated versus cocaine-associated cues. These responses were identified in the absence of the stimuli (cocaine or pups) and are proposed to be, in part, activation of these neurons related to motivational processing. Neither the distribution of neurons responding to pup-associated cue preference nor the demonstration that CART-expressing neurons are responsive to reward-associated cue preference has been previously reported. We hypothesize that the expression of preference for cocaine versus pup-associated cues is made possible by the concerted activity of these regionally distributed networks of neurons that are in part specific to the preference response.

Estrogen plus Progesterone Increases Progestin Receptor Immunoreactivity in the Brain of Ovariectomized Guinea Pigs

The goal of these experiments was to determine the number and distribution of brain cells that contain progestin receptors (PR) and to determine the effect of estrogen and estrogen plus progesterone on PR content of those cells. Ovariectomized adult female guinea pigs were treated with oil (control), or estrogen followed by oil, or estrogen followed by progesterone. As expected, only those animals treated with estrogen plus progesterone became sexually receptive. The cellular content of PR was determined using a monoclonal antibody to the receptor, and standard immunocytochemical techniques. Analysis of the PR-immunoreactive (PR-IR) cells consisted of: (1) mapping the anatomical distribution of PR-IR cells; (2) analyzing the effect of steroid hormones on PR-IR cell number, and (3) determining the effect of steroid hormones on PR immunoreactivity per cell. PR immunoreactivity was located exclusively in the nuclei of cells in the preoptic area and hypothalamus. The most dense collections of PR-IR cells were found in the preoptic area, ventrolateral nucleus of the hypothalamus, and infundibular nucleus. Estrogen caused a dramatic increase in the number of PR-IR cells in these cell groups. Sequential treatment with estrogen plus progesterone further increased PR-IR cell number, in the preoptic area by 65%, in the ventrolateral nucleus by 38%, and in the infundibular nucleus by 49%. A cell-by-cell rating of the PR immunoreactivity was carried out in these three cell groups. We found that the staining intensity across the populations of PR-IR cells was increased by estrogen and further increased by sequential estrogen plus progesterone. Alterations in cellular PR content may contribute importantly to the ability of progesterone target cell groups to perform their specialized roles in steroid-regulated activity.

Functional mapping of the neural circuitry of rat maternal motivation: effects of site-specific transient neural inactivation

The present review focuses on recent studies from our laboratory examining the neural circuitry subserving rat maternal motivation across postpartum. We employed a site‐specific neural inactivation method by infusion of bupivacaine to map the maternal motivation circuitry using two complementary behavioural approaches: unconditioned maternal responsiveness and choice of pup‐ over cocaine‐conditioned incentives in a concurrent pup/cocaine choice conditioned place preference task. Our findings revealed that, during the early postpartum period, distinct brain structures, including the medial preoptic area, ventral tegmental area and medial prefrontal cortex infralimbic and anterior cingulate subregions, contribute a pup‐specific bias to the motivational circuitry. As the postpartum period progresses and the pups grow older, it is further revealed that maternal responsiveness becomes progressively less dependent on the medial preoptic area and medial prefrontal cortex infralimbic activity, and more distributed in the maternal circuitry, such that additional network components, including the medial prefrontal cortex prelimbic subregion, are recruited with maternal experience, and contribute to the expression of late postpartum maternal behaviour. Collectively, our findings provide strong evidence that the remarkable ability of postpartum females to successfully care for their developing infants is subserved by a distributed neural network that carries out efficient and dynamic processing of complex, constantly changing incoming environmental and pup‐related stimuli, ultimately allowing the progression of appropriate expression and waning of maternal responsiveness across the postpartum period.

Microinfusion of cocaine into the medial preoptic area or nucleus accumbens transiently impairs maternal behavior in the rat.

Cocaine was microinfused bilaterally (50 microg/0.5 microl/side) into the medial preoptic area (MPOA) or nucleus accumbens (NA), 2 regions within the rat brain neural circuit known to mediate maternal behavior (MB). Additionally, 2 sites not involved in this neural circuit, the dorsal striatum and dorsal medial hippocampus, were used as control sites. Microinfusion of cocaine into the MPOA or NA impaired MB, whereas infusion into the control sites did not. MB impairment was not temporally coincident with the increased locomotor activity, also documented after cocaine infusion into the MPOA or NA, arguing strongly that impaired MB is a direct, specific effect of cocaine in these areas, not a derivative of increased motor activity. This is the first demonstration that cocaine action on single central nervous system (CNS) sites can impair MB to the same extent as systemic injections. Thus, cocaines simultaneous effect on multiple CNS sites is not required for MB impairment.

EVIDENCE FOR ESTROGEN RECEPTOR IN CELL NUCLEI AND AXON TERMINALS WITHIN THE LATERAL HABENULA OF THE RAT : REGULATION DURING PREGNANCY

The habenular complex is involved in several estrogen‐dependent reproductive behaviors in female rats, namely, sexual behavior, maternal behavior, and postpartum sexual behavior. Although it is known that estrogen acts in other brain regions to mediate these behaviors, it is not known whether estrogen may also act directly on the habenular complex. To address this possibility, we examined this region for the presence of estrogen receptor (ER). This analysis was carried out in separate experiments by using in situ hybridization, immunocytochemistry at the light and electron microscopic levels, and steroid autoradiography. Neurons within the lateral habenula (LHb), but not the medial habenula, express ER mRNA, contain ER immunoreactivity (ER‐ir) in their nuclei, and concentrate radiolabelled estradiol, providing strong evidence for the presence of functional ER in the lateral habenula. There were also ER‐ir containing punctate fibers within the LHb, which, at the electron microscopic level, in part, proved to be axons and presynaptic axonal terminals. Both the level of ER‐ir in cell nuclei and the density of ER‐ir fibers within the LHb were regulated during the course of pregnancy and the postpartum period, suggesting that the sensitivity of the LHb to estrogen may be altered during this time. Taken together, these results demonstrate that the LHb is likely a more estrogen‐sensitive region than was previously considered, and they suggest alternative mechanisms of action for ER. ER within the LHb may play a critical role in the involvement of the LHb in estrogen‐dependent female reproductive behaviors. J. Comp. Neurol. 392:330–342, 1998.

Lateral habenula neurons are necessary for the hormonal onset of maternal behavior and for the display of postpartum estrus in naturally parturient female rats.

In 16-day pregnant, hysterectomized-ovariectomized and estradiol benzoate-treated rats, cytotoxic lesions of the lateral habenula (Lhb) produced severe deficits in maternal behavior (K.P. Corodimas, J.S. Rosenblatt, M.E. Canfield, & J.I. Morrell, 1993). To determine if deficits could be found in parturient rats, females were bilaterally injected with kainic acid (KA) to produce cytotoxic lesions of the Lhb. Controls either received bilateral KA-induced lesions to the hippocampus or were unoperated. All females maintained their pregnancies, underwent parturition, and were tested for maternal behavior (pup retrieval, nursing, and nestbuilding), general activity, and oromotor carrying. Females with lesions of the Lhb had severe disruptions of all components of maternal behavior and postpartum estrus. These results extend previous findings that the Lhb is involved in the hormonal onset of maternal behavior and also demonstrate that the Lhb supports the display of postpartum estrus.

Induction of c-fos-like and fosB-like immunoreactivity reveals forebrain neuronal populations involved differentially in pup-mediated maternal behavior in juvenile and adult rats.

Juvenile rats can exhibit maternal behavior after being exposed continuously to rat pups, a process called sensitization. Maternal behavior in juveniles is robust and is similar to adult maternal behavior (Mayer and Rosenblatt [1979] Dev. Psychobiol. 12:407–424; Gray and Chesley [684] J. Comp. Psychol. 98:91–99). In this study, immunocytochemical detection of the protein products of two immediate‐early genes, c‐fos and fosB, was used as a tool to identify forebrain neuronal populations involved in the maternal behavior of 27‐day‐old juvenile rats compared with 60‐day‐old adults. To sensitize them, rats were exposed continuously to foster pups. Once they were maternal, they were isolated from pups overnight, reexposed to pups for 2 hours, and then killed. Nonmaternal control animals also were isolated overnight and were either reexposed to pups for 2 hours or kept isolated from pups before killing. The lateral habenula (LH) was the only area in which both maternal juveniles and maternal adults had more c‐Fos‐immunoreactive (‐Ir) neurons compared with controls. In maternal adults, the number of neurons that expressed c‐Fos and FosB immunoreactivity increased in the medial preoptic area (MPO) and the ventral bed nucleus of the stria terminalis (BSTv), whereas the dorsal bed nucleus of the stria terminalis (BSTd) and the medial and cortical nuclei of the amygdala (MEA and COA, respectively) had increases only in the number of neurons that expressed c‐Fos immunoreactivity. In contrast, juveniles, whether or not they were maternal, had the same number of c‐Fos‐IR and FosB‐Ir neurons in all these areas. The adult‐like increase in the number of c‐Fos‐Ir neurons found in maternal juveniles suggests that the juvenile LH participates in the neural circuit that supports maternal behavior in an adult‐like manner. The lack of c‐fos or fosB induction in the MPO, BSTv, BSTd, COA, or MEA of maternal juveniles compared with maternal adults may reflect the immaturity of these brain regions in juvenile rats. Exactly what this immaturity consists of and when the responses of these regions become adult‐like remain to be determined. J. Comp. Neurol. 416:45–78, 2000.

Circulating gonadal steroid hormones regulate estrogen receptor mRNA in the male rat forebrain

In male rats, the conversion of testosterone to estrogen via aromatization is a critical step in a number of androgen-mediated functions, especially reproductive behavior. Within the central nervous system (CNS), locally formed estrogen binds to its cognate estrogen receptor protein. Little is known about what factors regulate the expression of estrogen receptors in the male rat CNS. This study examined whether circulating male gonadal steroid hormones have a role in the regulation of estrogen receptor mRNA in brain regions critical for the expression of male reproductive behavior. Male rats were gonadectomized or sham operated, and 3 days later were sacrificed. Their brains were fixed by perfusion, frozen, and sectioned. Tissue sections were hybridized to an 35S-labeled 850 base cDNA probe, complementary primarily to the steroid binding domain of the estrogen receptor mRNA. Following post-hybridization washes, slides were dipped in photographic emulsion and exposed for 2 weeks. Estrogen receptor mRNA-containing neurons were observed in all brain regions previously shown by steroid hormone autoradiography to concentrate estrogen. Gonadectomy did not alter the number of estrogen receptor mRNA-producing neurons, but did produce a two-fold increase in the relative amount of estrogen receptor mRNA per cell in the medial preoptic nucleus, periventricular preoptic area, and bed nucleus of the stria terminalis. This study shows that circulating gonadal steroids down-regulate steady state levels of estrogen receptor mRNA within specific brain regions, and thereby have the potential to regulate the sensitivity of particular target regions in the CNS to estrogen.