Xigeng Zheng

Roles of BDNF, dopamine D(3) receptors, and their interactions in the expression of morphine-induced context-specific locomotor sensitization.

Drug seeking, craving, and relapse can be triggered by environmental stimuli that acquire motivational salience through repeated associations with the drugs effects. Previous studies indicated that the dopamine D(3) receptor (Drd3) might be involved in the expression of drug-conditioned responses in rats, and brain-derived neurotrophic factor (BDNF) could modulate Drd3 expression in the nucleus accumbens (NAc). However, the involvement of neural regions with Drd3 activation and the underlying interaction between BDNF and Drd3 in the expression of behavioral responses controlled by a drug-associated environment have remained poorly understood. The present study used a conditioning procedure to assess the roles of BDNF, Drd3, and their interactions in the NAc in the expression of morphine-induced context-specific locomotor sensitization. We showed that the expression of locomotor sensitization in the morphine-paired environment was accompanied by significantly increased expression of Drd3 mRNA and BDNF mRNA and protein levels. Both sensitized locomotion in morphine-paired rats and enhanced Drd3 mRNA were suppressed by intra-NAc infusion of anti-tyrosine kinase receptor B (TrkB) IgG. Furthermore, intra-NAc infusion of the Drd3-selective antagonist SB-277011A significantly decreased the expression of context-specific locomotor sensitization and upregulated BDNF mRNA. Altogether, these results suggest that BDNF/TrkB signaling and activation of Drd3 in the NAc are required for the expression of morphine-induced context-specific locomotor sensitization.

Coexistence of Anhedonia and anxiety-independent increased novelty-seeking behavior in the chronic mild stress model of depression

Previous research demonstrated excessive decreases in reward sensitivity and increases in harm avoidance in depressed individuals. These results straightly lead to a hypothesis that depressed patients should avoid novelty or express reduced novelty-seeking behavior. Nevertheless, literature in this regard is inconsistent. Furthermore, whether the potentially altered novelty-associated behavior is dependent on changed anxiety/fear or related to altered goal-directed approaching tendency is unclear. Here, we tested novel object-approaching behavior in a free-exploration paradigm in chronic mild stress (CMS)-induced anhedonic and stress-resistant rats respectively. Other CMS-induced, emotional behaviors were also examined in a battery of behavioral tests including novel cage, exploration, locomotor activity and elevated plus maze (EPM). We found that compared with controls, stress-resistant rats who consistently showed lower anxiety level in EPM (time in open arms) and, open-field (OF) test (time in central area) showed no sign of enhanced novel object approaching behavior. To the contrary, the anhedonic ones who did not express any sign of reduced anxiety showed paradoxically intensified novelty-approaching behavior. We concluded that reduced anxiety would not necessarily lead to enhanced novelty-seeking behavior; anhedonia coexists with anxiety-independent, increased novelty-seeking behavior. The salient paradox of coexistence of anhedonia and increased novelty-seeking behavior was critically discussed.

Orexin A attenuates unconditioned sexual motivation in male rats

Orexins are neuropeptides involved in multiple neurophysiological functions such as reward and motivation. However, it is not clear whether orexins are implicated in sexual motivation. This study aims to evaluate the effects of orexin A and the OX(1)R antagonist SB334867 on unconditioned sexual motivation. Forty-five male Wistar rats are divided into four groups. The four groups are respectively administered intracerebroventricularly with saline, orexin A (1, 10 microg), 10% DMSO (cyclodextrin) and SB334867 (5, 15 microg) 10-15 min before sexual motivation tests. The preference for a receptive female to a male in an open arena with two tethered animals is designated as unconditioned sexual motivation. The results show that orexin A reduces the female preference (reducing time in the female zone and/or increasing time in the male zone), the number of visits for the female zone and the total distance traveled in sexually high-motivated males. SB334867 has no effect on the female preference, the number of visits and the distance traveled in either sexually high-motivated or low-motivated males. Our experiments reveal that centrally administered orexin A attenuates sexual motivation in high-motivated males although endogenous orexin A might not play an important role in the expression of unconditioned sexual motivation.

Prefrontal high-frequency stimulation prevents sub-conditioning procedure-provoked, but not acute stress-provoked, reemergence of extinguished fear

We have recently shown that post-extinction exposure of rats to a sub-conditioning procedure (SCP, i.e., retraining with a shock intensity that is too weak to induce by itself significant fear conditioning) or to acute stress provokes reemergence of extinguished fear. Furthermore, this SCP effect can be abolished by high-frequency stimulation (HFS) of the medial prefrontal cortex (mPFC), when applied following the SCP. The aim of the present study was to test whether HFS of the mPFC is effective in preventing both SCP-induced and acute stress-provoked fear reemergence. Rats implanted with stimulating electrodes in the mPFC were trained to acquire high levels of freezing to conditioned auditory cue. This fear response was then extinguished. Three weeks later, no spontaneous recovery was observed, but rats exposed to either the SCP or acute stress again exhibited high levels of freezing. HFS of the mPFC, applied before provoking fear reemergence, prevented the effects of SCP, but not acute stress. These data suggest that acute stress may have more impact on functions of the mPFC and/or associated structures than a situational reminder of fear conditioning.

N-methyl-D-aspartate receptor-mediated glutamate transmission in nucleus accumbens plays a more important role than that in dorsal striatum in cognitive flexibility.

Cognitive flexibility is a critical ability for adapting to an ever-changing environment in humans and animals. Deficits in cognitive flexibility are observed in most schizophrenia patients. Previous studies reported that the medial prefrontal cortex-to-ventral striatum and orbital frontal cortex-to-dorsal striatum circuits play important roles in extra- and intra-dimensional strategy switching, respectively. However, the precise function of striatal subregions in flexible behaviors is still unclear. N-methyl-D-aspartate receptors (NMDARs) are major glutamate receptors in the striatum that receive glutamatergic projections from the frontal cortex. The membrane insertion of Ca2+-permeable α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptors (AMPARs) depends on NMDAR activation and is required in learning and memory processes. In the present study, we measured set-shifting and reversal learning performance in operant chambers in rats and assessed the effects of blocking NMDARs and Ca2+-permeable AMPARs in striatal subregions on behavioral flexibility. The blockade of NMDARs in the nucleus accumbens (NAc) core by AP5 impaired set-shifting ability by causing a failure to modify prior learning. The suppression of NMDAR-mediated transmission in the NAc shell induced a deficit in set-shifting by disrupting the learning and maintenance of novel strategies. During reversal learning, infusions of AP5 into the NAc shell and core impaired the ability to learn and maintain new strategies. However, behavioral flexibility was not significantly affected by blocking NMDARs in the dorsal striatum. We also found that the blockade of Ca2+-permeable AMPARs by NASPM in any subregion of the striatum did not affect strategy switching. These findings suggest that NMDAR-mediated glutamate transmission in the NAc contributes more to cognitive execution compared with the dorsal striatum.

Gene expression of conditioned locomotion and context-specific locomotor sensitization controlled by morphine-associated environment

The nucleus accumbens (NAc) is involved in contextual drug associations, which might be particularly important for environmental cue-induced relapse to drug seeking. In the present study, rats were first administered repeated morphine for 5 days (5 mg/kg, i.p.) in a contextually paired and unpaired design. After reexposure to the morphine-associated environment, which induced conditioned locomotor activity in the morphine-paired group, we performed a rat 27k 70-mer oligo array to profile gene expression in the NAc. One hundred fifty-five upregulated and 88 downregulated genes were found in the paired group compared with the unpaired group. Eight gene transcripts were then selected to confirm their alterations by quantitative real-time polymerase chain reaction (qRT-PCR). The identified genes generally play important roles in neuroactive receptor-ligand interactions, synapse plasticity, ion transport, and protein phosphorylation. Furthermore, the expression of the eight selected genes that were identified and confirmed to show significant fold changes in the first microarray experiment were again measured with qRT-PCR after morphine challenge (2 mg/kg, i.p.). As expected, 2 mg/kg morphine-induced context-specific sensitization. Meanwhile, mRNA expression of the selected genes showed marked upregulation in the morphine-paired group compared with the unpaired and acute groups. These results suggest that alterations in the expression of the identified genes in the NAc may contribute to the neuroplasticity underlying contextual cue-induced relapse to drug use.

Altered phosphorylation of GluA1 in the striatum is associated with locomotor sensitization induced by exposure to increasing doses of morphine

Escalation of drug consumption is involved in the transition from drug use to addiction. Our previous study demonstrated that neuronal activation in ventral tegmental area (VTA) and substantia nigra (SN) was associated with behavioral sensitization induced by increasing doses of morphine. Here we sought to characterize the molecular mechanism underlying this behavioral sensitization. We compared mitogen-activated protein kinase (MAPK) signaling following pretreatment with either increasing doses or fixed doses of morphine before and after behavioral sensitization. We found phospho-MAPK markedly increased in ventral striatum and decreased in dorsal striatum after either pretreatment group, but no further change after sensitization induced by 10mg/kg morphine challenge. Furthermore, we also evaluated the level of phospho-glutamate receptor 1 at serine 845 (pSer845-GluA1) and 831 (pSer831-GluA1) sites in ventral striatum and dorsal striatum. The results showed a significant increase in pSer845-GluA1/GluA1 ratio in ventral striatum but not dorsal striatum after pretreatment with increasing doses of morphine but not after fixed-dose or saline pretreatment. Importantly, pSer845-GluA1/GluA1 ratio was increased exclusively in dorsal striatum and not ventral striatum following acute morphine challenge specifically paired with increasing-dose pretreatment and not fixed-dose or saline. These findings indicate that behavioral sensitization-induced by chronic pretreatment with increasing doses of morphine might be more closely associated with the dynamic GluA1 activity in the striatum rather than the modulation of MAPK signaling. These findings also indicate that GluA1 phosphorylation may occur independent of MAPK activation.

Differential patterns of neuronal activation in rostral versus caudal ventral tegmental area involved in behavioral sensitization induced by an escalating-dose morphine administration paradigm

The transition to addiction often involves a gradual process of escalated drug intake. The purpose of the present study was to characterize neuronal activation in the ventral tegmental area (VTA) and substantia nigra (SN) following chronic escalating-dose morphine exposure (days 1-7, 2 mg/kg/d; days 8-21, beginning at 10 mg/kg/d, increasing by 2 mg/kg/d), with steady-dose morphine (2 mg/kg/d, i.p., for 21 days) as the comparison. Using immunohistochemical double-staining for tyrosine hydroxylase (TH) and Fos, we found that the number of Fos(+)TH(+) neurons in the rostral VTA and number of Fos(+)TH(-) neurons in the lateral SNr were significantly increased in escalating-dose morphine-treated rats compared with steady-dose morphine-treated rats and acute morphine-treated rats. Meanwhile, this increase was associated with robust expression of behavioral sensitization after a challenge with 10 mg/kg morphine. The number of Fos(+)TH(+) neurons was significantly increased by acute morphine in the caudal VTA and SNc, but this number did not increase further with morphine pretreatment. These results demonstrate that behavioral sensitization was associated with elevated activation of dopaminergic neurons in the rostral VTA and nondopaminergic neurons in the lateral SNr, which could only be induced by chronic escalating-dose morphine rather than chronic steady-dose morphine pretreatment.

Region-specific roles of the prelimbic cortex, the dorsal CA1, the ventral DG and ventral CA1 of the hippocampus in the fear return evoked by a sub-conditioning procedure in rats

The return of learned fear is an important issue in anxiety disorder research since an analogous process may contribute to long-term fear maintenance or clinical relapse. A number of studies demonstrate that mPFC and hippocampus are important in the modulation of post-extinction re-expression of fear memory. However, the region-specific role of these structures in the fear return evoked by a sub-threshold conditioning (SC) is not known. In the present experiments, we first examined specific roles of the prelimbic cortex (PL), the dorsal hippocampus (DH, the dorsal CA1 area in particular), the ventral hippocampus (the ventral dentate gyrus (vDG) and the ventral CA1 area in particular) in this fear return process. Then we examined the role of connections between PL and vCA1 with this behavioral approach. Rats were subjected to five tone-shock pairings (1.0-mA shock) to induce conditioned fear (freezing), followed by three fear extinction sessions (25 tone-alone trials each session). After a post-test for extinction memory, some rats were retrained with the SC procedure to reinstate tone-evoked freezing. Rat groups were injected with low doses of the GABAA agonist muscimol to selectively inactivate PL, DH, vDG, or vCA1 120 min before the fear return test. A disconnection paradigm with ipsilateral or contralateral muscimol injection of the PL and the vCA1 was used to examine the role of this pathway in the fear return. We found that transient inactivation of these areas significantly impaired fear return (freezing): inactivation of the prelimbic cortex blocked SC-evoked fear return in particular but did not influence fear expression in general; inactivation of the DH area impaired fear return, but had no effect on the extinction retrieval process; both ventral DG and ventral CA1 are required for the return of extinguished fear whereas only ventral DG is required for the extinction retrieval. These findings suggest that PL, DH, vDG, and vCA1 all contribute to the fear return and connections between PL and vCA1 may be involved in the modulation of this process.

High-dose corticosterone after fear conditioning selectively suppresses fear renewal by reducing anxiety-like response.

Exposure therapy is widely used to treat anxiety disorders, including posttraumatic stress disorder (PTSD). However, preventing the return of fear is still a major challenge after this behavioral treatment. An increasing number of studies suggest that high-dose glucocorticoid treatment immediately after trauma can alleviate the symptoms of PTSD in humans. Unknown is whether high-dose glucocorticoid treatment following fear conditioning suppresses the return of fear. In the present study, a typical fear renewal paradigm (AAB) was used, in which the fear response to an auditory cue can be restored in a novel context (context B) when both training and extinction occur in the same context (context A). We trained rats for auditory fear conditioning and administered corticosterone (CORT; 5 and 25mg/kg, i.p.) or vehicle with different delays (1 and 24h). Forty-eight hours after drug injection, extinction was conducted with no drug in the training context, followed by a test of tone-induced freezing behavior in the same (AAA) or a shifted (AAB) context. Both immediate and delayed administration of high-dose CORT after fear conditioning reduced fear renewal. To examine the anxiolytic effect of CORT, independent rats were trained for cued or contextual fear conditioning, followed by an injection of CORT (5 and 25mg/kg, i.p.) or vehicle at a 1 or 24h delay. One week later, anxiety-like behavior was assessed in the elevated plus maze (EPM) before and after fear expression. We found that high-dose CORT decreased anxiety-like behavior without changing tone- or context-induced freezing. These findings indicate that a single high-dose CORT administration given after fear conditioning may selectively suppress fear renewal by reducing anxiety-like behavior and not by altering the consolidation, retrieval, or extinction of fear memory.