Open access peer-reviewed chapter

Corticotrophin-Releasing Factor (CRF) through CRF1 Receptor Facilitates the Expression of Morphine-Related Positive and Aversive Memory in Mice

Written By

Pilar Almela, Juan A. García-Carmona, Elena Martínez-Laorden, María V. Milanés and María L. Laorden

Submitted: 05 May 2017 Reviewed: 25 July 2018 Published: 09 November 2018

DOI: 10.5772/intechopen.80504

From the Edited Volume

Opioids - From Analgesic Use to Addiction

Edited by Pilar Almela Rojo

Chapter metrics overview

1,144 Chapter Downloads

View Full Metrics

Abstract

Different studies have elucidated the mechanisms underlying the formation and expression of drug-related cue memories; corticotrophin-releasing factor (CRF) plays a critical role in reward- and aversion-driven associative learning. In the present chapter, we have evaluated whether CP-154,526, a selective CRF1 receptor (CRF1R) antagonist, or genetic deletion of CRF1R (KO mice) have comparable effects on conditioned place preference (CPP) and conditioned place aversion (CPA) learning. We also investigated CP-154,526 effects on morphine-induced CPP activation of CRF, CREB phosphorylation, and thioredoxin (Trx1) expression in dentate gyrus (DG), a brain region involved in memory consolidation, and the role of hypothalamic-pituitary-adrenocortical (HPA) axis in CPA expression and extinction. The CRF1R antagonist abolished the acquisition of morphine CPP, Trx-1 and BDNF increased expression, and pCREB/Trx-1 co-localization in the DG. The increase in adrenocorticotropic hormone (ACTH) plasma levels observed after CPA expression was attenuated in CRF1R KO mice, suggesting a role of HPA axis in aversive memories. Altogether, these results suggest a critical role of CRF, through CRF1R, in molecular changes involved in memory formation and consolidation and may facilitate the development of effective treatments for opioid addiction.

Keywords

  • conditioned place preference
  • conditioned place aversion
  • morphine
  • hippocampus
  • CRF
  • HPA axis

1. Introduction

Drug addiction is a chronic brain disease with a high rate of relapse [1, 2, 3]. Despite years of abstinence from drugs, relapse can occur when addicts encounter cues, including people or places, associated with their prior drug use [4]. Drug-associated memory can persist throughout the lifetime of a patient; therefore, the elimination of this kind of memory is considered to be crucial for the treatment of drug addiction.

In organism and human models, drug reward can be assessed using a Pavlovian conditioning procedure known as conditioned place preference/conditioned place aversion (CPP/CPA) [5, 6, 7]. CPP for the drug-paired environment is predicted by self-reported measures of drug liking in humans [6]. CPA for the drug-paired environment is used to infer the dysphoric properties of drugs, including opioid receptor antagonists [8]. Many neurotransmitters, neurotrophic factors, and protein kinases have been delineated in the regulation of the formation and expression of drug-associated reward memories and withdrawal-associated aversive memories [9, 10, 11, 12, 13].

Corticotrophin-releasing factor (CRF) in the brain plays a critical role in reward- and aversion-driven associative learning. However, it is not clear whether it does this by a common mechanism or by separated mechanisms that can be dissociated. The knowledge of these mechanisms could lead to more effective treatments for addictive processes. CRF and its CRF1 receptor (CRF1R) are widely distributed and in a highly conserved way in several brain regions, including the hippocampal formation, involved in reward reinforcement, craving and aversive effects of drug of abuse [14, 15, 16, 17]. At the extrahypothalamic level, CRF acts as a neuroregulator of the behavioral and emotional integration of environmental and endogenous stimuli associated with drug dependence [18, 19]. In the hippocampal dentate gyrus (DG), an important brain region involved in saving similar experiences and contexts [20], CRF is released from inhibitory interneurons [21] through CRF1R [14] by environmental signals. CRF1R activation stimulates Gαs protein, promoting the induction of the protein kinase A/cAMP response element binding protein (CREB) pathway [22]. CREB activity in the brain is critical for learning and memory processes [23], and it has been reported to be involved in the expression of opioid dependence. The activation of CREB, as one of the main downstream effectors of extracellular signal-regulated kinase (ERK), accelerates the transcription of CREB-dependent genes such as the brain-derived neurotrophic factor (BDNF). With respect to hypothalamus, CRF release from paraventricular nucleus (PVN) controls the hypothalamic-pituitary-adrenal (HPA) axis responses to stress and drug addiction [24, 25, 26]. CRF neurons in the PVN and CRF fibber in DG have direct connexion with dopaminergic neurons located in the ventral tegmental area (VTA) projecting to nucleus accumbens (NAc) [27, 28].

Advertisement

2. Role of CRF in the rewarding effects of morphine

CPP is an animal model widely used to evaluate the correlation between contexts and drugs. Different substances of abuse display differential ability to produce CPP. Opiates induce strong CPP over a wide range of experimental conditions [5]. Previous studies from our laboratory [29, 30, 31, 32] and others [33, 34] have demonstrated that morphine administration evokes significant CPP for the drug-associated environment. Different neurobiological substrates have been involved in the rewarding properties of drugs of abuse, although the mesolimbic dopaminergic pathway has been pointed out to be the critical system for drug reward. Recently, it has been suggested that PVN may have a role in the reinforcing effects of opioids [35]. Various studies have elucidated the mechanisms underlying the formation and expression of drug-related cue memories. CRF in the brain plays a critical role in reward-driven associative learning. During the formation or consolidation process (CPP expression), the majority of the CRF-positive neurons in the PVN, central nucleus of amygdale (CeA), and bed nucleus of stria terminalis (BNST) coexpresses pCREB after morphine-induced CPP, suggesting that drug-paired context could trigger neuronal activity in the brain stress system [29]. Morphine-treated mice in their home cage do not show any changes in total CRF/CREB positive neurons, indicating that the exposure to drug-paired environments is necessary for CRF activation in the brain stress system [29]. Anatomical and functional studies reveal connections between CRF and the mesolimbic dopaminergic system. Thus, VTA and NAc receive CRF-positive projections from the PVN and stress extrahypothalamic areas [36, 37], which have been proposed to regulate dopamine release. The rewarding effect of morphine (CPP expression) is decreased by pretreatment with CP-154,526, a selective CRF1 antagonist, suggesting an important role of CRF/CRF1 receptor in memory formation and consolidation [30].

2.1Implications of different signaling pathways in the rewarding effects of morphine. Role of CRF1 receptors

Hippocampus is a brain region known to participate in associative processes such as declarative memory, and PVN is an important stress area. Both structures are related with mesolimbic pathways [38]. Our group has studied the implication of different signaling pathways in both areas, because the understanding of how the formation of drug-reward memories alters the neurobiology of the hippocampal DG and PVN, and may shed light on the later and more persistent aspect of addiction.

The transcription factor CREB is critical in the conversion from short-term to long-term memory, and it is involved in the creation of long-term memory. Learning and memory and drug addiction share certain intracellular signaling pathways and depend on activation of CREB [39]. According to previous studies [40, 41], our laboratory has demonstrated that the number of pCREB positive neurons in PVN and DG is significantly increased after morphine-induced CPP expression (Figure 1). Since CRF1R is coupled to stimulatory G protein Gαs and can thus activate PKA and, subsequently, CREB [22], our group has investigated if CRF1R signaling is involved in CREB activity after morphine-induced CPP. Administration of the CRF1R antagonist, CP-154,526, completely revoked pCREB positive neuron enhancement induced by morphine in PVN and slightly in DG. CREB involvement in morphine dependence has been previously supported by studies demonstrating that CREB mutant mice do not respond to the reinforcing properties of morphine in a conditioned place preference paradigm [42], suggesting that specific CREB functions are necessary for the rewarding properties of this drug.

Figure 1.

CREB activation in PVN (A) and DG (C) after morphine-induced CPP. Scale bar 100 μm. Quantitative analysis of pCREB immunodetection in PVN (B) and DG (D). Data are expressed as mean ± SEM. **p < 0.01, ***p < 0.001 versus vehicle (veh) + saline (S); +p < 0.05, +++p < 0.001 versus veh + morphine (M). CP-154,526 (CP). Optical density (OD).

Although it is known that CRF signaling is involved in the drug withdrawal-induced anxiogenic-like and negative behavioral response [43], no definitive data are available about the role in the positive reinforcing properties of opiates. CRF-immunoreactive fibers densely innervate many intrahypothalamic and extrahypothalamic brain areas, such as hippocampus. Besides, CRF, through CRF1R, increases neuronal activity propagation from DG, the classical hippocampal input region, to the hypothalamic structure CA1 [44]. CRF is present in GABAergic hippocampal neurons of the pyramidal cells [14]. The supramammillary (SuM) region of the hypothalamus acts a connection nucleus between limbic and hypothalamic structures involved in controlling cognitive aspects [45]. Thus, SuM sends robust and direct inputs to DG. For example, it has been shown that mild stress could activate the SuM cells that project to the hippocampus [46]. Our group has previously shown that most of the CRF positive neurons in PVN coexpresses pCREB during morphine CPP. In addition, we have observed an enhancement in CRF fibers density in DG after morphine administration. Both changes were antagonized by injection of CP-154,526 (Figure 2). CRF binding to CRF1R results in activation of heterotrimeric G-proteins. The physiological functions of CRF1R in the central nervous system and in the periphery have been mainly associated to an increase in intracellular cAMP levels. This is consistent with a predominant coupling to Gαs (cAMP/PKA/CREB). However, CRF through CRFR1 is capable of activating other Gα types such as Gαs and activate inositol triphosphate (IP3) cascade. An enhancement in the concentration of secondary messengers (cAMP, IP3, and Ca2+) in cells, induced by CRF1R agonists, promotes the activation of several transcriptional factors such as CREB, AP-1, NF-κB, and the calcium response element (CARE) [47, 48, 49, 50, 51, 52, 53]. In this sense, the antagonist of the CRF1R, CP-154,526, by blocking the postsynaptic CRF1R, inhibited CREB phosphorylation in PVN and DG. Moreover, morphine treatment induced an increase in CRF fiber immunodetection in DG, suggesting an elevated CRF release, which was prevented by pretreatment with this antagonist. Since CRF1R activation increases Ca2+ levels, it is possible that CP-154,526 inhibits CRF release by blocking presynaptic CRF1R in PVN.

Figure 2.

CRF/pCREB double-labeling photomicrographs in PVN (A). The upper right side of the figure shows the quantitative analysis of double-labeled neurons (B). CRF fiber photomicrographs in the DG (C). The down right side of the figure shows the CRF fiber density in the DG (D). Scale bar 100 or 50 μm. Data are expressed as mean ± SEM. ***p < 0.001 versus vehicle (veh) + saline (S); ++p < 0.01, +++p < 0.001 versus veh + morphine (M). CP-154,526 (CP). Optical density (OD).

Several evidences suggest that CREB phosphorylation represents a site of convergence for various signaling pathways and alters gene expression [40]. CREB activation can also be regulated by the family of the redox protein Trx-1 [54]. In addition to its antioxidant activity, Trx-1 has been shown to play a crucial role in cellular signaling by controlling several important members of the signal transduction pathway. Thus, NF-κB, p38 mitogen-activated protein kinases, activator protein-1, CREB (as mentioned before), estrogen receptor, glucocorticoid receptor, and p53 are the targets of Trx-1 [55]. Data from our laboratory have shown that morphine-induced CPP increases Trx-1 expression in DG (Figure 3). Trx-1 might activate CREB phosphorylation, thus increasing the rewarding effects of morphine. In agreement with our results, other studies have also observed an increased Trx-1 expression following morphine or methamphetamine administration [56]. Upregulation of CREB activity induced by methamphetamine was suppressed by Trx-1siRNA, which suggests that Trx-1 is necessary for CREB activation [55, 56]. Moreover, morphine-induced Trx-1 expression is blocked by naloxone, indicating that morphine induces Trx-1 expression via activating opioid receptors [57]. Results from our laboratory showing a positive relationship between morphine rewarding effects, and Trx-1 expression are in contrast with another study [58] demonstrating that geranylgeranylacetone induces Trx-1 and, concomitantly, reduces morphine-induced CPP. These variations could be explained by the differential regulating roles of NAc and hippocampus. Besides, CREB expression has been shown to be increased in hippocampus but decreased in NAc after morphine conditioning [40], which suggests that CREB activity is differently regulated depending on the brain area studied. Our investigations have demonstrated a large number of pCREB/Trx-1 double-labeled neurons in DG (Figure 3). These neuron colocalizations in DG suggest that CREB might be activated by Trx-1 in this brain nucleus involved in memory consolidation processes. Due to the important role of TRX-1 in regulating the cellular redox balance, the induction of TRX-1 expression following morphine CPP could be associated to a mechanism of neural protection against a stressful situation.

Figure 3.

Characterization of pCREB and Trx-1 immunostaining in the dentate gyrus (DG) after morphine-induced CPP. (A) Schematic illustration showing the analyzed region of the DG (diagram modified from Franklin & Paxinos) [59]. Coordinate −1.94 mm from Bregma. (B) High-magnification image of a mouse midbrain coronal section immunostained for pCREB and Trx-1. Scale bar 100 μm. Representative confocal images of pCREB (red) (C–F) and Trx-1 (green) (C′–F′). Colocalization (pCREB/Trx-1) is shown in C″–F″ by yellow-orange neurons in the merged images. Scale bar 20 μm. Graphs on the right indicate the mean total number of pCREB (G), Trx-1 (H), and double-labeled (pCREB/Trx-1) neurons (I). Data are expressed as mean ± SEM. ***p < 0.001 versus vehicle (veh) + saline (S); +p < 0.05, ++p < 0.01 versus veh + morphine (M). CP-154,526 (CP).

Pretreatment with CP-154,526 completely blocks morphine-induced CPP elevation of Trx-1 expression in DG (Figure 3).

We have also shown an increase in the number of pCREB neurons coexpressing Trx-1 following morphine-induced CPP, so CRF1R could be involved in CREB phosphorylation, probably through a Trx-1-dependent way. The exact mechanism by which the CRF system participates in Trx-1 signaling regulation in DG is not completely understood. One possible explanation could indicate that pCREB binds to CRE in the 5′-upstream sequence of Trx-1 gene, thus inducing Trx-1 expression to regulate its phosphorylation. In agreement with this hypothesis, other authors have demonstrated that ephedrine promotes Trx-1 expression via the β-adrenergic receptor/cyclic AMP/PKA/DARPP-32 signaling pathway [60]. Besides, methamphetamine-induced CREB activity in rat pheochromocytoma cells was shown to be regulated by Trx-1 [56].

As shown in Figure 4, morphine-induced CPP increases the number of pCREB-positive neurons in PVN, an increase that was blocked by CP-154,526 treatment. However, there are no changes in the number of Trx-1 positive neurons or in the double labeled neurons (pCREB/Trx-1).

Figure 4.

Characterization of pCREB and Trx-1 immunostaining in the paraventricular nucleus (PVN) after morphine-induced CPP. (A) Schematic illustration showing the analyzed region of the PVN (diagram modified from Franklin & Paxinos) [59]. Coordinate −0.82 mm from Bregma. (B) High-magnification image of a mouse midbrain coronal section immunostained for pCREB and Trx-1. Scale bar 100 μm. Representative confocal images of pCREB (red) (C–F) and Trx-1 (green) (C′–F′). Colocalization (pCREB/Trx-1) is shown in C″–F″ by yellow-orange neurons in the merged images. Scale bar 20 μm. Graphs on the right indicate the mean total number of pCREB (G), Trx-1 (H), and double-labeled (pCREB/Trx-1) neurons (I). Data are expressed as mean ± SEM. **p < 0.01, versus vehicle (veh) + saline (S); ++p < 0.01, versus veh + morphine (M). CP-154,526 (CP).

On the other hand, BDNF, an important neurotrophin for synaptic plasticity, is one of the molecular candidates underlying the development of persistent neuroplastic adaptation that regulates drug addiction [61]. Several lines of evidence indicate that chronic morphine treatment triggers ERK activation in different brain regions [62]. ERK phosphorylates CREB and active (phosphorylated) CREB stimulates the expression of target genes, including BDNF [63, 64, 65]. Chronic morphine use has been shown to increase the expression of BDNF in the NAc and hippocampus [61, 66, 67]. According to these data, our findings demonstrated that morphine-induced CPP activates BDNF signaling in the DG without any changes in the saline group (Figure 5), demonstrating that repeated morphine with context exposure, but not merely the context, increases BDNF expression in DG, suggesting that BDNF is implicated in drug-induced contextual memory formation. Therefore, BDNF is a crucial signal molecule involved in morphine dependence. However, whether this molecule is regulated in a CRF1R-dependent manner remains largely unknown: CP-154,526 attenuated CREB-BDNF expression (Figures 4 and 5) and prevented morphine-induced CPP [29]. Taken together, CRF1R-mediated CREB-BDNF signaling changes may regulate morphine reward through modulating contextual memory in the hippocampus.

Figure 5.

Western-blotting analysis of BDNF in the dentate gyrus (DG) and paraventricular nucleus (PVN) from animals pretreated with vehicle (veh) or CP-154,526 (CP) before saline or morphine. The immunoreactivity corresponding to BDNF is expressed as a percentage of that in the control group defined as 100% value. ***p < 0.001 versus morphine + CP; +p < 0.05 versus saline + veh.

Advertisement

3. Role of CRF1 receptor in the aversive effects induced by naloxone-precipitated withdrawal

The physical component of morphine withdrawal syndrome can be assessed by scoring some somatic withdrawal signs after morphine exposure [68]. Recent results from our group have demonstrated significant alterations in some morphine withdrawal signs such as body weight loss, rearing, rubbing, grooming, diarrhea, freezing, and time to first immobility in wild type morphine-withdrawn animals compared with controls treated with saline (Figure 6). Besides, and in agreement with previous studies [69, 70, 71], our laboratory has shown that body weight loss (Figure 6H), freezing (Figure 6F), and diarrhea (Figure 6E) are significantly attenuated in CRF1R KO mice although an increase in jumping in CRF1R KO mice was observed (Figure 6A), as it has been described previously by other authors [72]. Jumping is a sensitive and commonly used index of naloxone-induced withdrawal [73, 74, 75, 76]. However, it is important to clarify that different neural elements mediate several withdrawal behaviors [77, 78]. Thus, it is not easy to extrapolate naloxone-precipitated jumping in CRF1R KO mice to other physical symptoms like body weight loss.

Figure 6.

Behavior effects by naloxone (nx)-precipitated morphine withdrawal in wild type (WT) or knockout (CRF1R KO) mice. The following somatic signs, (A) jumping, (B) rearing, (C) rubbing, (D) grooming, (E) diarrhea, (F) freezing behavior, and (H) body weight loss, induced after nx (1 mg/kg, s.c.)-injection to morphine or saline-treated mice during 18 min, were evaluated. The time to first immobilization (G) was also evaluated. Data are expressed as the mean ± SEM. $$p < 0.01 versus WT mice treated with morphine + nx; **p < 0.01 versus WT mice treated with saline + nx; +p < 0.05, ++p < 0.01, +++p < 0.001 versus WT mice treated with saline+nx; ##p < 0.01, ###p < 0.001 versus KO mice treated with saline + nx; &&&p < 0.001 versus WT mice treated with morphine + nx.

Advertisement

4. Role of CRF1 receptor in CPA expression and extinction

It is commonly accepted that affective drug withdrawal symptoms are of major motivational significance in contributing to relapse and continued drug use; thus, it is important to understand the mechanisms that mediate affective behaviors during morphine withdrawal. CPA paradigm is a highly sensitive animal model for the measurement of the negative affective component of drug withdrawal as well as to investigate the neural substrates underlying the aversive memory associated with drug withdrawal [79, 80]. In this model, a morphine-dependent animal undergoing withdrawal is exposed to a particular environment for a period of time. When later is given the opportunity to freely explore the apparatus, animals trained in this way tend to avoid the previously paired context due to the association between the context and aversive memories of drug withdrawal [79].

The extinction of this aversion occurs if the association is weakened by repeated exposure to the withdrawal-associated context in the absence of the conditioned stimulus, and the initial response (CPA) can be reinstated by a drug priming injection, stress or by conditioned cues. Extinction is complete when animals no longer avoid the previously cue-paired compartment. Typically, while memory reconsolidation requires single context reexposure, extinction requires multiple cue reexposures [81]. For example, fear conditioning studies suggest that the extinction process does not eliminate the initial context, but the organism learns that this cue does not cause the previous stimulus [82]. Thus, extinction requires associative learning, consolidation, and the formation of a new memory [83].

Recently, our group has investigated the mechanism underlying CPA expression and extinction. These experiments showed that morphine administration induced a significant place aversion for the naloxone-paired compartment, compared to the saline group. However, CRF1R KO mice presented less aversion than wild type mice (Figure 7A).

Figure 7.

(A) CPA expression induced by naloxone (nx, 1 mg/kg, s.c.) in wild type (WT) or knockout (CRF1R KO) mice treated with morphine or saline. The score was calculated for each mouse as the difference between the postconditioning and the preconditioning time spent in the naloxone-paired compartment. (B) Extinction of CPA training. Aversion scores from day 5 to 13 for WT and CRF1R KO mice are shown. Data are expressed as the mean ± SEM. +++p < 0.001 versus WT mice treated with saline + nx, &p < 0.05, &&p < 0.01, &&&p < 0.001 versus WT mice treated with morphine + nx.

There is much information about the neurobiological mechanisms involving extinction of reward memory of drug taking [84, 85, 86]. However, little information is known about extinction of aversive memory of drug withdrawal [87]. Previous studies have demonstrated that the aversive effects of opiates might be related to basal genotype differences in the brain systems [88]. Accordingly, we have clearly demonstrated that the genetic disruption of the CRF/CRF1R pathway decreases the period of CPA extinction (Figure 7B).

Thus, results obtained by our laboratory regarding CPA expression and extinction suggest an important role for CRF1R in aversive memory.

Advertisement

5. Role of HHA axis in the CPA induced by morphine withdrawal

It is well established that acute withdrawal of all major drugs of abuse dysregulates the HPA axis and alters CRF activity in the PVN of the hypothalamus, with a common response of increased adrenocorticotropic hormone (ACTH) and corticosterone [89], which mediate somatic and aversive components of withdrawal [72, 90, 91, 92]. To evaluate whether a causal link exists between CRF1R activation and HPA axis, our group has measured plasma ACTH and corticosterone levels in wild type and CRF1R KO mice after naloxone-induced CPA expression and CPA extinction (Figure 8). Our investigations have shown that plasma ACTH levels are increased in wild type mice although plasma corticosterone levels are not changed following CPA expression. These results indicate that ACTH-independent mechanisms could have an important role in the regulation of the adrenal stress system to appropriately adapt its response to physiological necessities, and even the presence of pituitary ACTH is basic for adrenocortical function. Numerous lines of evidence indicate that a large number of neuropeptides, neurotransmitters, growth factors, and bacterial ligands can influence the release of adrenal glucocorticoids independently of pituitary ACTH [93]. Adrenocortical cells express a large diversity of receptors for these factors, thus triggering potential direct actions on glucocorticoids release. Damage in the upstream stress regulating pathways in the brain leads to a rupture between ACTH and corticosterone, which suggests that central nervous system neurocircuits can regulate HPA axis response at both pituitary and adrenal sites [94]. Our results also indicate that CPA expression-induced ACTH release is attenuated in CRF1R KO mice. In agreement with these observations, it has been reported fewer ACTH levels in morphine withdrawn animals treated with CRF1R antagonists [70]. Besides, a role for the HPA axis and extra-hypothalamic brain circuitry in somatic, molecular, and endocrine changes induced during opioid withdrawal has been described [72]. ACTH plasma levels returned to basal in wild type and CRF1R KO mice after CPA extinction. These results suggest that CPA expression is, at least, partially due to an increase in plasma ACTH levels which can be decreased after naloxone CPA extinction.

Figure 8.

Effect of CPA expression and CPA extinction training on ACTH (A and B) and corticosterone (C and D) plasma levels in wild type (WT) and knockout (CRF1R KO) mice. Data are expressed as the mean ± SEM. +++p < 0.001 versus WT mice treated with saline + nx, &&&p < 0.001 versus WT mice treated with morphine + nx.

Advertisement

6. Conclusion

CP-154,526 administration or genetic deletion of CRF1R impairs CPP and CPA learning, suggesting that the expression of reward and aversive learning and memory shares some common neural circuits related with CRF/CRF1R signaling. During the formation or consolidation process (CPP expression), the majority of phospho-CREB positive neurons in DG coexpresses Trx-1, in parallel with an increased expression of BDNF, suggesting that Trx-1 could activate CREB and this in turn accelerates the transcription of CREB-dependent genes such as BDNF. However, CP-154,526 diminishes CPP expression, in parallel with a block of phospho-CREB/Trx-1 colocalization and BDNF expression, suggesting that Trx-1-CREB-BDNF signaling could be essential for memory formation or consolidation. In addition, CPA expression training increases plasma ACTH levels, which is critical for the maintenance of aversive memories associated with drug withdrawal. Genetic deletion of CRF1R (KO mice) induces a reduction in CPA expression accompanied with a higher decrease in ACTH plasma levels. CPA extinction period is reduced in KO mice, indicating a role for CRF1R in the aversive memory retrieval. Altogether, these results indicate a critical role for CRF, through CRF1R, in molecular changes involved in reward memory-associated behaviors and in aversive memory expression and extinction. The disruption of these processes by CRF1 antagonists might lead to effective treatments in drug addiction.

Advertisement

Acknowledgments

This research was supported by a grant from the Ministerio de Economía, Industria y Competitividad (SAF2017-85679-R).

References

  1. 1. Leshner AI. Drug abuse and addiction treatment research. The next generation. Archives of General Psychiatry. 1997;54:691-694
  2. 2. O’Brien CP, Childress AR, McLellan AT, Ehrman R. Classical conditioning in drug-dependent humans. Annals of the New York Academy of Sciences. 1992;654:400-415
  3. 3. O’Brien CP, McLellan AT. Myths about the treatment of addiction. Lancet. 1996;347:237-240
  4. 4. Childers A, Ehrman R, McLellan AT, O’Brien C. Conditioned craving and arousal in cocaine addiction: A preliminary report. NIDA Research Monograph. 1988;81:74-80
  5. 5. Tzschentke TM. Measuring reward with the conditioned place preference (CPP) paradigm: Update of the last decade. Addiction Biology. 2007;12:227-462. DOI: 10.1111/j.1369-1600.2007.00070.x
  6. 6. Childs E, de Wit H. Amphetamine-induced place preference in humans. Biological Psychiatry. 2009;65:900-904. DOI: 10.1016/j.biopsych.2008.11.016
  7. 7. Childs E, de Wit H. Contextual conditioning enhances the psychostimulant and incentive properties of d-amphetamine in humans. Addiction Biology. 2013;18:985-992. DOI: 10.1111/j.1369-1600.2011.00416.x
  8. 8. Martin del Campo AF, McMurray RG, Besser GM, Grossman A. Effect of 12-hour infusion of naloxone on mood and cognition in normal male volunteers. Biological Psychiatry. 1992;32:344-353. DOI: 10.1016/0006-3223(92)90038-2
  9. 9. LaLumiere RT, Kalivas PW. Glutamate release in the nucleus accumbens core is necessary for heroin seeking. The Journal of Neuroscience. 2008;28:3170-3177. DOI: 10.1523/JNEUROSCI.5129-07
  10. 10. Lee AM, Messing RO. Protein kinases and addiction. Annals of the New York Academy of Sciences. 2008;1141:22-57. DOI: 10.1196/annals.1441.022
  11. 11. Ron D, Jurd R. The ‘ups and downs’ of signaling cascades in addiction. Science’s STKE. 2005;2005:re14. DOI: 10.1126/stke.3092005re14
  12. 12. Russo SJ, Mazei-Robison MS, Ables JL, Nestler EJ. Neurotrophic factors and structural plasticity in addiction. Neuropharmacology. 2009;56(Suppl 1):73-82. DOI: 10.1016/j.neuropharm.2008.06.059
  13. 13. Wise RA. Dopamine and reward: The anhedonia hypothesis 30 years on. Neurotoxicity Research. 2008;14:169-183. DOI: 10.1007/BF03033808
  14. 14. Chen Y, Bender RA, Brunson KL, Pomper JK, Grigoriadis DE, Wurst W, et al. Modulation of dendritic differentiation by corticotropin-releasing factor in the developing hippocampus. PNAS. 2004;101:15782-15787. DOI: 10.1073/pnas.0403975101
  15. 15. Korosi A, Veening JG, Kozicz T, Henckens M, Dederen J, Groenink L. Distribution and expression of CRF receptor 1 and 2 mRNAs in the CRF over-expressing mouse brain. Brain Research. 2006;1072:46-55. DOI: 10.1016/j.brainres.2005.12.034
  16. 16. Lim MM, Tsivkovskaia NO, Bai Y, Young LJ, Ryabinin AE. Distribution of corticotropin-releasing factor and urocortin 1 in the vole brain. Brain, Behavior and Evolution. 2006;68:229-240. DOI: 10.1159/000094360
  17. 17. Refojo D, Echenique C, Muller MB, Reul JM, Deussing JM, Wurst W, et al. Corticotropin-releasing hormone activates ERK1/2MAPK in specific brain areas. PNAS. 2005;102:6183-6188. DOI: 10.1073/pnas.0502070102
  18. 18. García-Carmona JA, Almela P, Baroja-Mazo A, Milanés MV, Laorden ML. Restricted role of CRF1R for the activity of brainstem catecholaminergic neurons in the negative state of morphine withdrawal. Psychopharmacology. 2012;220:379-393. DOI: 10.1007/s00213-011-2478-y
  19. 19. Haass-Koffler CL, Bartlett SE. Stress and addiction: Contribution of the corticotropin releasing factor (CRF) system in neuroplasticity. Frontiers in Molecular Neuroscience. 2012;5:1-13. DOI: 10.3389/fnmol.2012.00091
  20. 20. Vivar C, van Praag H. Functional circuits of new neurons in the dentate gyrus. Neural Circuits. 2013;7:1-13. DOI: 10.3389/fncir.2013.00015
  21. 21. Chen Y, Bender RA, Frotscer M, Baram TZ. Novel and transient populations of corticotropin-releasing hormone-expressing neurons in developing hippocampus suggest unique functional roles: A quantitative spatiotemporal analysis. The Journal of Neuroscience. 2001;21:7171-7181
  22. 22. Kasagi Y, Horiba N, Sakai K, Fukuda Y, Suda T. Involvement of cAMP-response element binding protein in corticotropin-releasing factor (CRF)-induced down-regulation of CRF receptor 1 gene expression in rat anterior pituitary cells. Journal of Neuroendocrinology. 2002;14:587-592
  23. 23. Briand LA, Lee BG, Lelay J, Kaestner KH, Blendy JA. Serine 133 phosphorylation is not required for hippocampal CREB-mediated transcription and behavior. Learning & Memory. 2015;22:109-115. DOI: 10.1101/lm.037044.114
  24. 24. Dallman MF, Akana SF, Strack AM, Hanson ES, Sebastian RJ. The neural network that regulates energy balance is responsive to glucocorticoids and insulin and also regulates HPA axis responsivity at a site proximal to CRF neurons. Annals of the New York Academy of Sciences. 1995;771:730-742
  25. 25. Martín F, Laorden ML, Milanés MV. Morphine withdrawal regulates phosphorylation of cAMP response element binding protein (CREB) through PKC in the nucleus tractus solitarius-A2 catecholaminergic neurons. Journal of Neurochemistry. 2009;110:1422-1432. DOI: 10.1111/j.1471-4159.2009.06234
  26. 26. Martín F, Mora L, Laorden ML, Milanés MV. Protein kinase C phosphorylates the cAMP response element binding protein in the hypothalamic paraventricular nucleus during morphine withdrawal. British Journal of Pharmacology. 2011;163:857-875. DOI: 10.1111/j.1476-5381.2011.01287.x
  27. 27. Voorn P, Jorritsma-Byham B, Van Dijk C, Buijs RM. The dopaminergic innervation of the ventral striatum in the rat: A light- and electron-microscopical study with antibodies against dopamine. The Journal of Comparative Neurology. 1986;251:84-99. DOI: 10.1002/cne.902510106
  28. 28. Séguéla P, Watkins KC, Descarries L. Ultrastructural features of dopamine axon terminals in the anteromedial and the suprarhinal cortex of adult rat. Brain Research. 1988;442:11-22
  29. 29. García-Carmona JA, Milanés MV, Laorden ML. Brain stress system response after morphine-conditioned place preference. The International Journal of Neuropsychopharmacology. 2013;10:1-13. DOI: 10.1017/S1461145713000588
  30. 30. García-Carmona JA, Camejo DM, Almela P, Jiménez A, Milanés MV, Sevilla F, Laorden ML. CP-154,526 modifies CREB phosphorylation and thioredoxin-1 expression in the dentate gyrus following morphine-induced conditioned place preference. PLoS One. 2015;10(8):e0136164. DOI: 10.1371/journal.pone.0136164
  31. 31. González-Cuello A, Mora L, Hidalgo JM, Meca N, Lasheras C, Milanés MV, Laorden ML. Enhanced tyrosine hydroxylase phosphorylation in the nucleus accumbens and nucleus tractus solitarius-A2 cell group after morphine-conditioned place preference. Naunyn-Schmiedeberg’s Archives of Pharmacology. 2010;382:525-534. DOI: 10.1007/s00210-010-0567-9
  32. 32. Lasheras MC, Laorden ML, Milanés MV, Núñez C. Corticotropin-releasing factor 1 receptor mediates the activity of the reward system evoked by morphine-induced conditioned place preference. Neuropharmacology. 2015;95:168-180. DOI: 10.1016/j.neuropharm.2014.12.021
  33. 33. Ma YY, Meng L, Guo CHY, Han JS, Lee D, Cui CL. Dose- and time-dependent, context-induced elevation of dopamine and its metabolites in the nucleus accumbens of morphine-induced CPP rats. Behavioural Brain Research. 2009;204:192-199. DOI: 10.1016/j.bbr.2009.06.017
  34. 34. Zarrindast MR, Ebrahimi-Ghiri M, Rostami P, Rezayof A. Repeated pre-exposure to morphine into the ventral pallidum enhances morphine-induced place preference: Involvement of dopaminergic and opioidergic mechanisms. Behavioural Brain Research. 2007;181:35-41. DOI: 10.1016/j.bbr.2007.03.019
  35. 35. Ewan EE, Martin TJ. Rewarding electrical brain stimulation in rats after peripheral nerve injury: Decreased facilitation by commonly abused prescription opioids. Anesthesiology. 2011;115:1271-1280. DOI: 10.1097/ALN.0b013e3182330448
  36. 36. Rodaros D, Caruana DA, Amir S, Stewart J. Corticotropin-releasing factor projections from limbic forebrain and paraventricular nucleus of the hypothalamus to the region of the ventral tegmental area. Neuroscience. 2007;105:8-13. DOI: 10.1016/j.neuroscience.2007.09.043
  37. 37. Chen YW, Rada PV, Bützler BP, Leibowitz SF, Hoebel BG. Corticotropin releasing factor in the nucleus accumbens shell induces swim depression, anxiety, and anhedonia along with changes in local dopamine/acetyl-choline balance. Neuroscience. 2012;206:155-166. DOI: 10.1016/j.neuroscience.2011.12.009
  38. 38. Ewan EE, Martin TJ. Analgesics as reinforcers with chronic pain: Evidence from operant studies. Neuroscience Letters. 2013;557(Pt A):60-64. DOI: 10.1016/j.neulet.2013.08.018
  39. 39. Nestler EJ. Common molecular and cellular substrates of addiction and memory. Neurobiology of Learning and Memory. 2002;73:637-647. DOI: 10.1006/nlme.2002.4084
  40. 40. Zhou LF, Zhu YP. Changes of CREB in rat hippocampus, prefrontal cortex and nucleus accumbens during three phases of morphine induced conditioned place preference in rats. Journal of Zhejiang University. Science. B. 2006;7:107-113. DOI: 10.1631/jzus.2006.B0107
  41. 41. Moron JA, Gullapalli S, Taylor C, Gupta A, Gomes I, Deve LA. Modulation of opiate-related signalling molecules in morphine-dependent conditioned behaviour: Conditioned place preference to morphine induces CREB phosphorylation. Neuropsychopharmacology. 2010;35:955-966. DOI: 10.1038/npp.2009.199
  42. 42. Walters CL, Blendy JA. Different requirements for cAMP response element binding protein in positive and negative reinforcing properties of drugs of abuse. The Journal of Neuroscience. 2001;21:9438-9444
  43. 43. Koob GF. A role of brain stress system in addiction. Neuron. 2008;59:11-34. DOI: 10.1016/j.neuron.2008.06.012
  44. 44. Von Wolff G, Avrabos CH, Stepan J, Wurst W, Deussing JM, Holsboer F, Eder M. Voltage-sensitive dye imaging demonstrated an enhancing effect of corticotropin-releasing hormone on neuronal activity propagation through the hippocampal formation. Journal of Psychiatric Research. 2011;45:256-261. DOI: 10.1016/j.jpsychires.2010.06.007
  45. 45. Gonzalo-Ruiz A, Morte L, Flecha JM, Sanz JM. Neurotransmitter characteristics of neurons projecting to the suprammamillary nucleus of the rat. Anatomy and Embryology (Berlin). 1999;200:377-392
  46. 46. Choi WK, Wirtshafter D, Park HJ, Lee MS, Her S, Shim I. The characteristics of supramammillary cells projecting to the hippocampus in stress response in rat. The Korean Journal of Physiology & Pharmacology. 2012;16:17-24. DOI: 10.4196/kjpp.2012.16.1.17
  47. 47. Wiesner B, Roloff B, Fechner K, Slominski A. Intracellular calcium measurements of single human skin cells after stimulation with corticotropin-releasing factor and urocortin using confocal laser scanning microscopy. Journal of Cell Science. 2003;116:1261-1268. DOI: 10.1242/jcs.00301
  48. 48. Fazal N, Slominski A, Choudhry MA, Wei ET, Sayeed MM. Effect of CRF and related peptides on calcium signaling in human and rodent melanoma cells. FEBS Letters. 1998;435:187-190. DOI: 10.1016/S0014-5793(98)01067-9
  49. 49. Pisarchik A, Slominski A. Molecular and functional characterization of novel CRFR1 isoforms from the skin. European Journal of Biochemistry. 2004;27:2821-2830. DOI: 10.1111/j.1432-1033.2004.04216.x
  50. 50. Zbytek B, Pfeffer LM, Slominski AT. Corticotropin-releasing hormone stimulates NF-kappaB in human epidermal keratinocytes. The Journal of Endocrinology. 2004;181:R1-R7
  51. 51. Zbytek B, Slominski AT. Corticotropin-releasing hormone induces keratinocyte differentiation in the adult human epidermis. Journal of Cellular Physiology. 2005;203:118-126. DOI: 10.1002/jcp.20209
  52. 52. Zmijewski MA, Slominski AT. CRF1 receptor splicing in epidermal keratinocytes: Potential biological role and environmental regulations. Journal of Cellular Physiology. 2009;218:593-602. DOI: 10.1002/jcp.21632
  53. 53. Inda C, Armando NG, Dos Santos Claro PA, Silberstein S. Endocrinology and the brain: Corticotropin-releasing hormone signalling. Endocrine Connections. 2017;6:R99-R120. DOI: 10.1530/EC-17-0111
  54. 54. Hirota K, Matsui M, Murata M, Takashima Y, Cheng FS, et al. Nucleoredoxin, glutaredoxin and thioredoxin differentially regulate NF-kB, AP-1 and CREB activation in HEK293 cells. Biochemical and Biophysical Research Communications. 2000;274:177-182. DOI: 10.1006/bbrc.2000.3106
  55. 55. Luo FC, Qi L, Lv T, Wang SD, Liu H, et al. Geranylgeranylacetone protects mice against morphine induced hyperlocomotion, rewarding effect, and withdrawal syndrome. Free Radical Biology & Medicine. 2012;52:1218-1227. DOI: 10.1016/j.freeradbiomed.2012.01.004
  56. 56. Lv T, Wang SD, Bai J. Thioredoxin-1 was required for CREB activity by methamphetamine in rat pheochromocytoma cells. Cellular and Molecular Neurobiology. 2013;33:319-325. DOI: 10.1007/s10571-012-9897-0
  57. 57. Luo FC, Feng YM, Zhao L, Kui L, Wang SD, Song JY, et al. Thioredoxin-1 expression regulated by morphine in sh-sy5y cells. Neuroscience Letters. 2012;523:50-55. DOI: 10.1016/j.neulet.2012.06.039
  58. 58. Kondo N, Nakamura H, Masutani H, Yodoi J. Redox regulation of human thioredoxin network. Antioxidants & Redox Signaling. 2006;8:1881-1890. DOI: 10.1089/ars.2006.8.1881
  59. 59. Franklin KBJ, Paxinos G. The Mouse Brain in Stereotaxic Coordinates. 3rd ed. San Diego: Academic Press; 2008. p. 256
  60. 60. Jia JJ, Zeng XS, Li Y, Ma S, Bai J. Ephedrine induced thioredoxin-1 expression through β-adrenergic receptor/cyclic AMP/protein kinase A/dopamine-and cyclic AMP-regulated phosphoprotein signaling pathway. Cellular Signalling. 2013;25:1194-1201. DOI: 10.1016/j.cellsig.2013.02.007
  61. 61. Zhong P, Liu Y, Hu Y, Wang T, Zhao YP, Liu QS. BDNF interacts with endocannabinoids to regulate cocaine-induced synaptic plasticity in mouse midbrain dopamine neurons. The Journal of Neuroscience. 2015;35:4469-4481. DOI: 10.1523/JNEUROSCI.2924-14
  62. 62. Lv XF, Sun LL, Cui CL, Han JS. Nac shell Arc/Arg3.1 protein mediates reconsolidation of morphine CPP by increased GluR1 cell surface expression: Activation of ERK-coupled CREB is required. The International Journal of Neuropsychopharmacology. 2015;18:pyv030. DOI: 10.1093/ijnp/pyv030
  63. 63. Kandel ER. The molecular biology of memory storage: A dialog between genes and synapses. Bioscience Reports. 2001;21:565-611. DOI: 10.1126/science.1067020
  64. 64. Barco A, Alarcon JM, Kandel ER. Expression of constitutively active CREB protein facilitates the late phase of long-term potentiation by enhancing synaptic capture. Cell. 2002;108:689-703. DOI: 10.1016/S0092-8674(02)00657-8
  65. 65. Bramham CR, Messaoudi E. BDNF function in adult synaptic plasticity: The synaptic consolidation hypothesis. Progress in Neurobiology. 2005;76:99-125. DOI: 10.1016/j.pneurobio.2005.06.003
  66. 66. Alvandi MS, Bourmpoula M, Homberg JR, Fathollahi Y. Association of contextual cues with morphine reward increases neural and synaptic plasticity in the ventral hippocampus of rats. Addiction Biology. 2017;22:1883-1894. DOI: 10.1111/adb.12547
  67. 67. Wan L, Xie Y, Su L, Liu Y, Wang Y, Wang Z. Rack1 affects morphine reward via BDNF. Brain Research. 2011;1416:26-34. DOI: 10.1016/j.brainres.2011.07.045
  68. 68. Radke AK, Hotz NA, Gewirtz ME, Carroll ME. Reduced emotional signs of opiate withdrawal in rats selectively bred for low (LoS) versus high (HiS) saccharin intake. Psychopharmacology. 2013;227:117-126. DOI: 10.1007/s00213-012-2945-0
  69. 69. García-Carmona JA, Almela P, Baroja-Mazo A, Milanés MV, Laorden ML. Restricted role of CRF1 receptor for the activity of brainstem catecholaminergic neurons in the negative state of morphine withdrawal. Psychopharmacology. 2012;220:379-393. DOI: 10.1007/s00213-011-2478-y
  70. 70. Navarro-Zaragoza J, Núñez C, Laorden ML, Milanés MV. Effects of corticotropin-releasing factor receptor-1 antagonists on the brain stress system response to morphine withdrawal. Molecular Pharmacology. 2010;77:864-873. DOI: 10.1124/mol.109.062463
  71. 71. Iredale PA, Alvaro JD, Lee Y, Terwilliger R, Chen YL, Duman RS. Role of corticotropin-releasing factor receptor-1 in opiate withdrawal. Journal of Neurochemistry. 2000;74:199-208. DOI: 10.1046/j.1471-4159.2000.0740199.x
  72. 72. Papaleo F, Kitchener P, Contarino A. Disruption of the CRF/CRF1 receptor stress system exacerbates the somatic signs of opiate withdrawal. Neuron. 2007;53:577-589. DOI: 10.1016/j.neuron.2007.01.022
  73. 73. Blum K, Eubanks JD, Wiggins B, Wallace JE. Morphine withdrawal reactions in male and female mice. The American Journal of Drug and Alcohol Abuse. 1976;3:363-368
  74. 74. El-Kadi AO, Sharif SI. The influence of various experimental conditions on the expression of naloxone-induced withdrawal symptoms in mice. General Pharmacology. 1994;25:1505-1510
  75. 75. Miyamoto Y, Takemori AE. Relative involvement of supraspinal and spinal mu opioid receptors in morphine dependence in mice. Life Sciences. 1993;52:1039-1044
  76. 76. Way EL, Lou HH, Shen FH. Simultaneous quantitative assessment of morphine tolerance and physical dependence. The Journal of Pharmacology and Experimental Therapeutics. 1969;167:1-8
  77. 77. Brandao ML. Involvement of opioid mechanisms in the dorsal periaqueductal gray in drug abuse. Reviews in the Neurosciences. 1993;4:397-405
  78. 78. Koob GF, Maldonado R, Stinus L. Neural substrates of opiate withdrawal. Trends in Neurosciences. 1992;15:186-191
  79. 79. Myers KM, Bechtholt-Gompf AJ, Coleman BR, Carlezon WA. Extinction of conditioned opiate withdrawal in rats in a two-chambered place conditioning apparatus. Nature Protocols. 2012;7:517-526. DOI: 10.1038/nprot.2011.458
  80. 80. Stinus L, Caille S, Koob GF. Opiate withdrawal-induced place aversion lasts for up to 16 weeks. Psychopharmacology. 2000;149:115-120
  81. 81. Power AE, Berlau DJ, McGaugh JL, Steward O. Anisomycin infused into the hippocampus fails to block “reconsolidation” but impairs extinction: The role of re-exposure duration. Learning & Memory. 2006;13:27-34. DOI: 10.1101/lm.91206
  82. 82. Bouton ME. Context and behavioral processes in extinction. Learning & Memory. 2004;11:485-494. DOI: 10.1101/lm.78804
  83. 83. Milad MR, Quirk GJ. Neurons in medial prefrontal cortex signal memory for fear extinction. Nature. 2002;420:70-74. DOI: 10.1038/nature01138
  84. 84. Feltenstein MW, See RE. NMDA receptor blockade in the basolateral amygdala disrupts consolidation of stimulus-reward memory and extinction learning during reinstatement of cocaine-seeking in an animal model of relapse. Neurobiology of Learning and Memory. 2007;88:435-444. DOI: 10.1016/j.nlm.2007.05.006
  85. 85. Hsu E, Packard MG. Medial prefrontal cortex infusions of bupivacaine or AP-5 block extinction of amphetamine conditioned place preference. Neurobiology of Learning and Memory. 2008;89:504-512. DOI: 10.1016/j.nlm.2007.08.006
  86. 86. Torregrossa MM, Sanchez H, Taylor JR. D-cycloserine reduces the context specificity of pavlovian extinction of cocaine cues through actions in the nucleus accumbens. The Journal of Neuroscience. 2010;30:10526-10533. DOI: 10.1523/JNEUROSCI.2523-10.2010
  87. 87. Myers KM, Carlezon WA Jr. D-cycloserine facilitates extinction of naloxone-induced conditioned place aversion in morphine-dependent rats. Biological Psychiatry. 2010;67:85-87. DOI: 10.1016/j.biopsych.2009.08.015
  88. 88. Solecki W, Turek A, Kubik J, Przewlocki R. Motivational effects of opiates in conditioned place preference and aversion paradigm-a study in three inbred strains of mice. Psychopharmacology. 2009;207:245-255. DOI: 10.1007/s00213-009-1672-7
  89. 89. Ueno K, Maeda T, Kiguchi N, Kobayashi Y, Ozaki M, Kishioka S. Availability of serum corticosterone level for quantitative evaluation of morphine withdrawal in mice. Drug Discoveries & Therapeutics. 2011;5:71-75. DOI: 10.5582/ddt.2011.v5.2.71
  90. 90. Contarino A, Papaleo F. The corticotropin-releasing factor receptor-1 pathway mediates the negative affective states of opiate withdrawal. Proceedings of the National Academy of Sciences of the United States of America. 2005;102:18649-18654. DOI: 10.1073/pnas.0506999102
  91. 91. Harris GC, Aston-Jones G. Activation in extended amygdale corresponds to alter hedonic processing during protracted morphine withdrawal. Behavioural Brain Research. 2007;176:251-258. DOI: 10.1016/j.bbr.2006.10.012
  92. 92. Koob GF. A role for brain stress system in addiction. Neuron. 2008;59:11-34. DOI: 10.1016/j.neuron.2008.06.012
  93. 93. Bornstein SR, Engeland WC, Ehrhart-Bornstein ME, Herman JP. Dissociation of ACTH and glucocorticoids. Trends in Endocrinology and Metabolism. 2008;19:175-180. DOI: 10.1016/j.tem.2008.01.009
  94. 94. Choi DC, Furai AR, Evanson NK, Ostrander MM, Ulrich-Lai YM, Herman JP. Bed nucleus of the stria terminalis subregions differentially regulates hypothalamic-pituitary-adrenal axis activity: Implications for the integration of limbic inputs. The Journal of Neuroscience. 2007;27:2025-2034. DOI: 10.1523/JNEUROSCI.4301-06.2007

Written By

Pilar Almela, Juan A. García-Carmona, Elena Martínez-Laorden, María V. Milanés and María L. Laorden

Submitted: 05 May 2017 Reviewed: 25 July 2018 Published: 09 November 2018