Impact of gender on corticotropin-releasing factor and noradrenergic sensitivity in cocaine use disorder.

Responses to stress may be important in understanding gender differences in substance use disorders and may also be a target for development of treatment interventions. A growing body of both preclinical and clinical research supports important underlying gender differences in the corticotropin-releasing factor (CRF) and noradrenergic systems, which may contribute to drug use. Preclinical models have demonstrated increased sensitivity of females to CRF and noradrenergic-induced drug reinstatement compared with males, and, consistent with these findings, human laboratory studies have demonstrated greater sensitivity to corticotropin-releasing hormone (CRH) and noradrenergic stimulation in cocaine-dependent women compared with men. Furthermore, neuroimaging studies have demonstrated increased neural response to stressful stimuli in cocaine-dependent women compared with men as well as showing significant sex differences in the sensitivity of brain regions responsible for regulating the response to CRH. Development of interventions targeting the noradrenergic system and stress response in drug-dependent individuals could have important clinical implications for both women and men. © 2016 Wiley Periodicals, Inc.

Role of Corticotropin Releasing Factor 1 Signaling in Cocaine Seeking during Early Extinction in Female and Male Rats.

Locus coeruleus norepinephrine (LC-NE) and corticotropin releasing factor (CRF) neurons are involved in stress responses, including stress's ability to drive drug relapse. Previous animal studies indicate that female rats exhibit greater drug seeking than male rats during initial drug abstinence. Moreover, females are more sensitive to the effect of stress to drive drug seeking than males. Finally, LC-NE neurons are more sensitive to CRF in females compared to males. We hypothesized that increased drug seeking in females on extinction day one (ED1) is due to increased response to the stress of early withdrawal and is dependent upon the increased response of LC in females to CRF. We predicted that LC-NE neurons would exhibit Fos activation on ED1, and that blocking CRF1 signaling would decrease drug seeking on ED1 measured by responding on an active lever previously associated with cocaine self- administration. After chronic cocaine self-administration, female and male rats underwent a test for initial extinction responding by measuring lever pressing in the absence of cocaine. Prior to this Extinction Day 1 (ED1) session, rats were injected with vehicle or the selective CRF1 antagonist (CP) to measure effects of CRF antagonism on drug seeking during early abstinence. ED1 increased corticosterone in female rats, in proportion to lever responding in male and female, indicating that ED1 was stressful. Pretreatment with CP decreased cocaine seeking on ED1 more effectively in female compared to male rats. This increase in responding was associated with an increase in activation of LC NE neurons. Together, these findings indicate that stress, and signaling at CRF receptors in LC, may be involved in the increased drug seeking during initial abstinence.

Role of orexin/hypocretin in conditioned sucrose-seeking in female rats.

The orexin/hypocretin system has recently been implicated in reward-seeking, especially for highly salient food and drug rewards. Given that eating disorders affect women more than men, we reasoned that the orexin system may be strongly engaged in female rats, and during periods of food restriction as we recently reported in male rats. Therefore, the present study examined the involvement of the orexin system in operant responding for sucrose, and in cue-induced reinstatement of extinguished sucrose-seeking, in ad libitum fed vs. food-restricted female subjects. Female Sprague Dawley rats were trained to self-administer sucrose pellets, and we determined the effects of pretreatment with the OxR1 receptor antagonist SB 334867 (SB; 10-30 mg/kg) on fixed ratio (FR) sucrose self-administration, and on cue-induced reinstatement of extinguished sucrose-seeking. SB decreased sucrose self-administration in food-restricted but not in ad libitum-fed females. SB did not alter active lever responding during cue-induced reinstatement of sucrose-seeking in either feeding group. These results confirm our previous results in male rats that signaling at the OxR1 receptor is involved in the sucrose reinforcement and self-administration in food-restricted subjects. However, the finding that SB is ineffective at attenuating cue-induced reinstatement in females, but was effective in food-restricted males, leads us to conclude that food seeking induced by conditioned stimuli engages the orexin system differentially in males and females.

Prior access to a sweet is more protective against cocaine self-administration in female rats than in male rats.

It is well established that female rats are more sensitive than male rats to the reinforcing effects of cocaine (Lynch, 2008 [42] for review). We hypothesized that greater preference for cocaine would support greater avoidance of a cocaine-paired taste cue in female vs. male rats. Moreover, at least in male rats, greater avoidance of the taste cue is associated with greater cocaine self-administration (Grigson and Twining, 2002 [3]). Thus, we anticipated that female rats would not only demonstrate greater avoidance of the drug-paired taste cue, but greater drug-taking as well. We tested these hypotheses by examining avoidance of a saccharin cue in male and female rats following several pairings with self-administered saline or cocaine (0.16, 0.33, or 0.66 mg/infusion). Contrary to expectations, the results showed that female rats exhibited less avoidance of the cocaine-associated saccharin cue than male rats and self-administered less, rather than more, cocaine, Thus, while female rats reportedly take more drug than male rats when the drug is presented in the absence of an alternative reward, they take less drug than male rats when the opportunity to self-administer cocaine is preceded by access to a palatable sweet. Females, then, may not simply be more sensitive to the rewarding properties of drug, but also to the reinforcing properties of natural rewards and this increase in sensitivity to sweets may serve to protect against drug-taking behavior.

Attenuation of saccharin-seeking in rats by orexin/hypocretin receptor 1 antagonist.

RATIONALE: The orexin (Orx)/hypocretin system has been implicated in reward-seeking, especially for highly salient food and drug rewards. We recently demonstrated that signaling at the OxR1 receptor is involved in sucrose reinforcement and reinstatement of sucrose-seeking elicited by sucrose-paired cues in food-restricted rats. Because sucrose reinforcement has both a hedonic and caloric component, it remains unknown what aspect of this reward drives its reinforcing value. OBJECTIVES: The present study examined the involvement of the Orx system in operant responding for saccharin, a noncaloric, hedonic (sweet) reward, and in cue-induced reinstatement of extinguished saccharin-seeking in ad libitum-fed vs food-restricted male subjects. METHODS: Male Sprague Dawley rats were fed ad libitum or food-restricted and trained to self-administer saccharin. We determined the effects of pretreatment with the OxR1 receptor antagonist SB-334867 (SB; 10-30 mg/kg) on fixed ratio (FR) saccharin self-administration and on cue-induced reinstatement of extinguished saccharin-seeking. RESULTS: SB decreased responding and number of reinforcers earned during FR responding for saccharin and decreased cue-induced reinstatement of extinguished saccharin-seeking. All of these effects were obtained similarly in food-restricted and ad libitum-fed rats. CONCLUSIONS: These results indicate that signaling at the OxR1 receptor is involved in saccharin reinforcement and reinstatement of saccharin-seeking elicited by saccharin-paired cues regardless of food restriction. These findings lead us to conclude that the Orx system contributes to the motivational effects of hedonic food rewards, independently of caloric value and homeostatic needs.

Role of orexin/hypocretin in conditioned sucrose-seeking in rats.

RATIONALE: The orexin/hypocretin system has recently been implicated in reward-seeking, especially for highly salient food and drug rewards. We reasoned that this system may be strongly engaged during periods of reward restriction, including food restriction. OBJECTIVES: This study examined the involvement of the orexin (Orx) system in responding for sucrose, and in cue-induced reinstatement of extinguished sucrose-seeking, in ad libitum fed versus food-restricted male subjects. METHODS: Sprague-Dawley rats (n = 108) were trained to self-administer sucrose, and we determined the effects of pretreatment with the OxR1 receptor antagonist SB-334867 (SB; 10-30 mg/kg) on fixed ratio (FR) or progressive ratio (PR) sucrose self-administration, as well as on cue-induced reinstatement of sucrose-seeking. Finally, expression of the immediate early gene c-fos in Orx neurons was examined after self-administration, late extinction or cue-induced reinstatement of sucrose seeking. RESULTS: SB decreased lever responding (by about 1/3) and the number of reinforcers earned during FR, and less so during PR, schedules and decreased cue-induced reinstatement to sucrose-seeking to extinction levels, predominately in food-restricted rats. Additionally, Fos expression in Orx neurons in perifornical and dorsomedial hypothalamus was increased during extinction. CONCLUSIONS: These results indicate that signaling at the OxR1 receptor is involved in pronounced sucrose reinforcement, and reinstatement of sucrose-seeking elicited by sucrose-paired cues, in food-restricted subjects. These findings lead us to conclude that conditioned activation of Orx neurons increases motivation for food reward during food restriction.

A history of bingeing on fat enhances cocaine seeking and taking.

Binge eating and substance dependence are disorders characterized by a loss of control over consummatory behaviors. Given the common characteristics of these two types of disorders, it is not surprising that the comorbidity between eating disorders and substance abuse disorders is high (20-40%; Conason et al., 2006). It is unknown, however, whether loss of control in one disorder predisposes an individual to loss of control in the other. The present study, therefore, used a rodent model to test whether a history of binge eating would augment subsequent responding for cocaine. Using the limited access protocol described by Corwin et al. (1998), 45 adult male Sprague-Dawley rats were maintained on one of four dietary protocols for a period of six weeks: chow only (Chow; n = 9), continuous access to an optional source of dietary fat (Ad Lib; n = 12), 1-h access to an optional source of dietary fat daily (Daily; n = 12), or 1-h access to an optional source of dietary fat on Monday, Wednesday, and Friday (MWF; n = 12). All four groups also had unrestricted access to a nutritionally complete diet of chow and water. Fat-bingeing behaviors developed in the MWF rats, the group with the most restricted access to the optional fat. Thereafter, cocaine-seeking and -taking behaviors were assessed in all rats using a self-administration protocol modified from that described by Deroche-Gamonet et al. (2004), which focused on the motivation for and preoccupation with obtaining and consuming drug (assessed using a progressive ratio [PR] schedule of reinforcement) and persistence in responding for drug during periods of signaled drug non-availability (SNA). Rats with the MWF history tended to take more cocaine late in fixed ratio (FR) training, they persisted in their efforts to obtain cocaine in the face of signaled non-availability, worked harder for cocaine on a PR schedule of reinforcement, and exhibited more goal-directed behavior toward the cocaine-associated operandum. These results demonstrate a link between binge-type intake of fat and the development of drug-seeking and -taking behaviors, suggesting that a history of fat bingeing may predispose individuals to exhibit more robust "addiction-like" behaviors toward a substance of abuse. Thus, it appears that conditions promoting excessive behavior toward one substance (e.g., a palatable fatty food) beget excessive behavior toward another (e.g., cocaine).

c-Fos induction by a 14 T magnetic field in visceral and vestibular relays of the female rat brainstem is modulated by estradiol.

There is increasing evidence that high magnetic fields interact with the vestibular system of humans and rodents. In rats, exposure to high magnetic fields of 7 T or above induces locomotor circling and leads to a conditioned taste aversion if paired with a novel taste. Sex differences in the behavioral responses to magnetic field exposure have been found, such that female rats show more locomotor circling and enhanced conditioned taste aversion compared to male rats. To determine if estrogen modulates the neural response to high magnetic fields, c-Fos expression after 14 T magnetic field exposure was compared in ovariectomized rats and ovariectomized rats with estradiol replacement. Compared to sham exposure, magnetic field exposure induced significantly more c-Fos positive cells in the nucleus of the solitary tract and the parabrachial, medial vestibular, prepositus, and supragenualis nuclei. Furthermore, there was a significant asymmetry in c-Fos induction between sides of the brainstem in several regions. In ovariectomized rats, there was more c-Fos expressed in the right side compared to left side in the locus coeruleus and parabrachial, superior vestibular, and supragenualis nuclei; less expression in the right compared to left side of the medial vestibular; and no asymmetry in the prepositus nucleus and the nucleus of the solitary tract. Chronic estradiol treatment modulated the neural response in some regions: less c-Fos was induced in the superior vestibular nucleus and locus coeruleus after estradiol replacement; estradiol treatment eliminated the asymmetry of c-Fos expression in the locus coeruleus and supragenualis nucleus, created an asymmetry in the prepositus nucleus and reversed the asymmetry in the parabrachial nucleus. These results suggest that ovarian steroids may mediate sex differences in the behavioral responses to magnetic field exposure at the level of visceral and vestibular nuclei of the brainstem.

Role of orexin/hypocretin in reward-seeking and addiction: implications for obesity.

Orexins (also named hypocretins) are recently discovered neuropeptides made exclusively in the hypothalamus. Recent studies have shown that orexin cells located specifically in lateral hypothalamus (LH) are involved in motivated behavior for drugs of abuse as well as natural rewards. Administration of orexin has been shown to stimulate food consumption, and orexin signaling in VTA has been implicated in intake of high-fat food. In self-administration studies, the orexin 1 receptor antagonist SB-334867 (SB) attenuated operant responding for high-fat pellets, sucrose pellets and ethanol, but not cocaine, demonstrating that signaling at orexin receptors is necessary for reinforcement of specific rewards. The orexin system is also implicated in associations between rewards and relevant stimuli. For example, Fos expression in LH orexin neurons varied in proportion to conditioned place preference (CPP) for food, morphine, or cocaine. This Fos expression was altered accordingly for CPP administered during protracted abstinence from morphine or cocaine, when preference for natural rewards was decreased and drug preference was increased. Additionally, orexin has been shown to be involved in reward-stimulus associations in the self-administration paradigm, where SB attenuated cue-induced reinstatement of extinguished sucrose- or cocaine-seeking. Although the specific circuitry mediating the effects of orexin on food reward remains unknown, VTA seems likely to be a critical target for at least some of these orexin actions. Thus, recent studies have established a role for orexin in reward-based feeding, and further investigation is warranted for determining whether function/dysfunction of the orexin system may contribute to the overeating associated with obesity. The paper represents an invited review by a symposium, award winner or keynote speaker at the Society for the Study of Ingestive Behavior [SSIB] Annual Meeting in Portland, July 2009.

Labyrinthectomy abolishes the behavioral and neural response of rats to a high-strength static magnetic field.

Vertigo is a commonly-reported side effect of exposure to the high magnetic fields found in magnetic resonance imaging machines. Although it has been hypothesized that high magnetic fields interact with the vestibular apparatus of the inner ear, there has been no direct evidence establishing its role in magnet-induced vertigo. Our laboratory has shown that following exposure to high magnetic fields, rats walk in circles, acquire a conditioned taste aversion (CTA), and express c-Fos in vestibular and visceral relays of the brainstem, consistent with vestibular stimulation and vertigo or motion sickness. To determine if the inner ear is required for these effects, rats were chemically labyrinthectomized with sodium arsanilate and tested for locomotor circling, CTA acquisition, and c-Fos induction after exposure within a 14.1 T magnet. Intact rats circled counterclockwise after 30-min exposure to 14.1 T, but labyrinthectomized rats showed no increase in circling after magnetic field exposure. After 3 pairings of 0.125% saccharin with 30-min exposure at 14.1 T, intact rats acquired a profound CTA that persisted for 14 days of extinction testing; labyrinthectomized rats, however, did not acquire a CTA and showed a high preference for saccharin similar to sham-exposed rats. Finally, significant c-Fos was induced in the brainstem of intact rats by 30-min exposure to 14.1 T, but magnetic field exposure did not elevate c-Fos in labyrinthectomized rats above sham-exposed levels. These results demonstrate that an intact inner ear is necessary for all the observed effects of exposure to high magnetic fields in rats.

Evidence for a cephalic site of action of high magnetic fields on the behavioral responses of rats.

Static high magnetic fields (MFs) from 7 T to 9 T can elicit behavioral responses in rodents such as suppression of rearing, locomotor circling, and acquisition of a conditioned taste aversion (CTA). MF exposure also induces c-Fos expression in the visceral and vestibular nuclei of the brainstem, suggesting the stimulation of some sensory pathways. It is not clear, however, if the effects of the MF are caused by exposure to the uniform maximal field at the center of the magnet, or by exposure to the steep field gradients along the bore of the magnet during the rat's placement. In addition, the site of action within the rat is unknown. In an attempt to limit MF exposure to rostral or caudal portions of the rats' body, we exposed male and female rats at different positions within the bore of a 14.1-T superconducting magnet ranging from 2 cm (1.6 T at the head) to 155 cm (0.05 T at the head), with the center of the bore at 65 cm (14.1 T across the whole body). This approach also allowed us to expose rats to the maximal field strength (14.1 T) vs. the maximal field gradients (54 T/m). To assess both immediate and delayed behavioral effects, locomotor and CTA responses were recorded. A small but significant CTA was seen after exposure of the head to the lowest MF tested (0.05 T at 155 cm). Graded effects were seen, however, with greater circling and CTA acquisition as the MF strength increased at the rostral end of the rat. This suggests a cephalic site of action. Furthermore, maximal circling and CTA were induced after exposure to the uniform center field, and not after exposure to high field gradients on either side of the center. This suggests that the behavioral responses seen after MF exposure are a consequence of the uniform static field at the center of the magnet, and are not caused by passage through, or exposure to, the vertical field gradients. Female rats responded similarly to male rats, although magnet-induced CTA appeared resistant to extinction in female rats.

Sex and estrous cycle differences in the behavioral effects of high-strength static magnetic fields: role of ovarian steroids.

Advances in magnetic resonance imaging are driving the development of higher-resolution machines equipped with high-strength static magnetic fields (MFs). The behavioral effects of high-strength MFs are largely uncharacterized, although in male rats, exposure to 7 T or above induces locomotor circling and leads to a conditioned taste avoidance (CTA) if paired with a novel taste. Here, the effects of MFs on male and female rats were compared to determine whether there are sex differences in behavioral responses and whether these can be explained by ovarian steroid status. Rats were given 10-min access to a novel saccharin solution and then restrained within a 14-T magnet for 30 min. Locomotor activity after exposure was scored for circling and rearing. CTA extinction was measured with two-bottle preference tests. In experiment 1, males were compared with females across the estrous cycle after a single MF exposure. Females circled more and acquired a more persistent CTA than males; circling was highest on the day of estrus. In experiment 2, the effects of three MF exposures were compared among intact rats, ovariectomized females, and ovariectomized females with steroid replacement. Compared with intact rats, ovariectomy increased circling; estrogen replacement blocked the increase. Males acquired a stronger initial CTA but extinguished faster than intact or ovariectomized females. Thus the locomotor circling induced by MF exposure was increased in females and modulated by ovarian steroids across the estrous cycle and by hormone replacement. Furthermore, female rats acquired a more persistent CTA than male rats, which was not dependent on estrous phase or endogenous ovarian steroids.

Estrous changes in vaginal nociception in a rat model of endometriosis.

A rat model of endometriosis, in which pieces of uterine horn (versus fat in controls) are autotransplanted into the abdomen where they form cysts, reduces fecundity and produces vaginal hyperalgesia. The cysts gradually enlarge over a 2-month period postsurgically and then plateau. Cysts regress with low estrogen levels and reappear when they rise. Based on the hypothesis that the vaginal hyperalgesia depends upon the cysts, this study tested two predictions: that (1) the hyperalgesia would develop postsurgically in parallel with the cysts, and (2) the hyperalgesia would vary with estrous, being greatest when estrogen levels are high (proestrus) and least when low (estrus). In rats trained to escape vaginal distention, percentage escape responses to different distention volumes were measured across the rat's 4-day estrous cycle for 2.5 months before and up to 4 months after autotransplantation of uterus (n=9) or fat (n=6) in abdominal sites. Vaginal pressures were also measured. In rats with uterine but not fat autotransplants, escape percentages increased postsurgically over a 2-month period and then plateaued. The increase was greatest in proestrus and failed to occur in estrus. Vaginal pressures were unchanged in all groups. These results strongly support the hypothesis that the vaginal hyperalgesia depends upon the cysts. Because the cysts were located in sites remote from the vagina, the hyperalgesia involves viscero-visceral interactions and is likely centrally mediated, whereas the estrous modulation could involve hormonal actions either on the cysts or, more likely, on vaginal afferent fibers, and/or on central neurons.