Cortical adrenoceptor expression, function and adaptation under conditions of cannabinoid receptor deletion.

A neurochemical target at which cannabinoids interact to have global effects on behavior is brain noradrenergic circuitry. Acute and repeated administration of a cannabinoid receptor synthetic agonist is capable of increasing multiple indices of noradrenergic activity. This includes cannabinoid-induced 1) increases in norepinephrine (NE) release in the medial prefrontal cortex (mPFC); 2) desensitization of cortical α2-adrenoceptor-mediated effects; 3) activation of c-Fos in brainstem locus coeruleus (LC) noradrenergic neurons; and 4) increases in anxiety-like behaviors. In the present study, we sought to examine adaptations in adrenoceptor expression and function under conditions of cannabinoid receptor type 1 (CB1r) deletion using knockout (KO) mice and compare these to wild type (WT) controls. Electrophysiological analysis of α2-adrenoceptor-mediated responses in mPFC slices in WT mice showed a clonidine-induced α2-adrenoceptor-mediated increase in mPFC cell excitability coupled with an increase in input resistance. In contrast, CB1r KO mice showed an α2-adrenoceptor-mediated decrease in mPFC cell excitability. We then examined protein expression levels of α2- and ß1-adrenoceptor subtypes in the mPFC as well as TH expression in the locus coeruleus (LC) of mice deficient in CB1r. Both α2- and ß1-adrenoceptors exhibited a significant decrease in expression levels in CB1r KO mice when compared to WT in the mPFC, while a significant increase in TH was observed in the LC. To better define whether the same cortical neurons express α2A-adrenoceptor and CB1r in mPFC, we utilized high-resolution immunoelectron microscopy. We localized α2A-adrenoceptors in a knock-in mouse that expressed a hemoagglutinin (HA) tag downstream of the α2A-adrenoceptor promoter. Although the α2A-adrenoceptor was often identified pre-synaptically, we observed co-localization of CB1r with α2-adrenoceptors post-synaptically in the same mPFC neurons. Finally, using receptor binding, we confirmed prior results showing that α2A-adrenoceptor is unchanged in mPFC following acute or chronic exposure to the synthetic cannabinoid receptor agonist, WIN 55,212-2, but is increased, following chronic treatment followed by a period of abstinence. Taken together, these data provide convergent lines of evidence indicating cannabinoid regulation of the cortical adrenergic system.

Gene knockout of 5-lipoxygenase rescues synaptic dysfunction and improves memory in the triple-transgenic model of Alzheimer's disease.

5-Lipoxygenase (5LO) is upregulated in Alzheimer's disease (AD) and in vivo modulates the amyloidotic phenotype of amyloid precursor protein transgenic mice. However, no data are available on the effects that 5LO has on synaptic function, integrity and cognition. To address this issue, we used a genetic and a pharmacological approach by generating 3 × Tg mice deficient for 5LO and administering 3 × Tg mice with a 5LO inhibitor. Compared with controls, we found that even before the development of overt neuropathology, both animals manifested significant memory improvement, rescue of their synaptic dysfunction and amelioration of synaptic integrity. In addition, later in life, these mice had a significant reduction of Aß and tau pathology. Our findings support a novel functional role for 5LO in regulating synaptic plasticity and memory. They establish this protein as a pleiotropic contributor to the development of the full spectrum of the AD phenotype, making it a valid therapeutic target for the treatment of AD.

Opiate exposure and withdrawal dynamically regulate mRNA expression in the serotonergic dorsal raphe nucleus.

Previous results from our lab suggest that hypofunctioning of the serotonergic (5-HT) dorsal raphe nucleus (DRN) is involved in stress-induced opiate reinstatement. To further investigate the effects of morphine dependence and withdrawal on the 5-HT DRN system, we measured gene expression at the level of mRNA in the DRN during a model of morphine dependence, withdrawal and post withdrawal stress exposure in rats. Morphine pellets were implanted for 72h and then either removed or animals were injected with naloxone to produce spontaneous or precipitated withdrawal, respectively. Animals exposed to these conditions exhibited withdrawal symptoms including weight loss, wet dog shakes and jumping behavior. Gene expression for brain-derived neurotrophic factor (BDNF), tyrosine kinase receptor B (TrkB), corticotrophin releasing-factor (CRF)-R1, CRF-R2, alpha 1 subunit of the GABAA receptor (GABAA-α1), µ-opioid receptor (MOR), 5-HT1A receptor, tryptophan hydroxylase2 (TPH2) and the 5-HT transporter was then measured using quantitative real-time polymerase chain reaction at multiple time-points across the model of morphine exposure, withdrawal and post withdrawal stress. Expression levels of BDNF, TrkB and CRF-R1 mRNA were decreased during both morphine exposure and following 7days of withdrawal. CRF-R2 mRNA expression was elevated after 7days of withdrawal. 5-HT1A receptor mRNA expression was decreased following 3h of morphine exposure, while TPH2 mRNA expression was decreased after 7days of withdrawal with swim stress. There were no changes in the expression of GABAA-α1, MOR or 5-HT transporter mRNA. Collectively these results suggest that alterations in neurotrophin support, CRF-dependent stress signaling, 5-HT synthesis and release may underlie 5-HT DRN hypofunction that can potentially lead to stress-induced opiate relapse.

Stress-induced sensitization of cortical adrenergic receptors following a history of cannabinoid exposure.

The cannabinoid receptor agonist, WIN 55,212-2, increases extracellular norepinephrine levels in the rat frontal cortex under basal conditions, likely via desensitization of inhibitory α2-adrenergic receptors located on norepinephrine terminals. Here, the effect of WIN 55,212-2 on stress-induced norepinephrine release was assessed in the medial prefrontal cortex (mPFC), in adult male Sprague-Dawley rats using in vivo microdialysis. Systemic administration of WIN 55,212-2 30 min prior to stressor exposure prevented stress-induced cortical norepinephrine release induced by a single exposure to swim when compared to vehicle. To further probe cortical cannabinoid-adrenergic interactions, postsynaptic α2-adrenergic receptor (AR)-mediated responses were assessed in mPFC pyramidal neurons using electrophysiological analysis in an in vitro cortical slice preparation. We confirm prior studies showing that clonidine increases cortical pyramidal cell excitability and that this was unaffected by exposure to acute stress. WIN 55,212-2, via bath application, blocked postsynaptic α2-AR mediated responses in cortical neurons irrespective of exposure to stress. Interestingly, stress exposure prevented the desensitization of α2-AR mediated responses produced by a history of cannabinoid exposure. Together, these data indicate the stress-dependent nature of cannabinoid interactions via both pre- and postsynaptic ARs. In summary, microdialysis data indicate that cannabinoids restrain stress-induced cortical NE efflux. Electrophysiology data indicate that cannabinoids also restrain cortical cell excitability under basal conditions; however, stress interferes with these CB1-α2 AR interactions, potentially contributing to over-activation of pyramidal neurons in mPFC. Overall, cannabinoids are protective of the NE system and cortical excitability but stress can derail this protective effect, potentially contributing to stress-related psychopathology. These data add to the growing evidence of complex, stress-dependent modulation of monoaminergic systems by cannabinoids and support the potential use of cannabinoids in the treatment of stress-induced noradrenergic dysfunction.

Morphine history sensitizes postsynaptic GABA receptors on dorsal raphe serotonin neurons in a stress-induced relapse model in rats.

The serotonin (5-hydroxytryptamine, 5-HT) system plays an important role in stress-related psychiatric disorders and substance abuse. Previous work has shown that the dorsal raphe nucleus (DR)-5-HT system is inhibited by swim stress via stimulation of GABA synaptic activity by the stress neurohormone corticotropin-releasing factor (CRF). Additionally, the DR 5-HT system is regulated by opioids. The present study tests the hypothesis that the DR 5-HT system regulates stress-induced opioid relapse. In the first experiment, electrophysiological recordings of GABA synaptic activity in 5-HT DR neurons were conducted in brain slices from Sprague-Dawley rats that were exposed to swim stress-induced reinstatement of previously extinguished morphine conditioned place preference (CPP). Behavioral data indicate that swim stress triggers reinstatement of morphine CPP. Electrophysiology data indicate that 5-HT neurons in the morphine-conditioned group exposed to stress had increased amplitude of inhibitory postsynaptic currents (IPSCs), which would indicate greater postsynaptic GABA receptor density and/or sensitivity, compared to saline controls exposed to stress. In the second experiment, rats were exposed to either morphine or saline CPP and extinction, and then 5-HT DR neurons from both groups were examined for sensitivity to CRF in vitro. CRF induced a greater inward current in 5-HT neurons from morphine-conditioned subjects compared to saline-conditioned subjects. These data indicate that morphine history sensitizes 5-HT DR neurons to the GABAergic inhibitory effects of stress as well as to some of the effects of CRF. These mechanisms may sensitize subjects with a morphine history to the dysphoric effects of stressors and ultimately confer an enhanced vulnerability to stress-induced opioid relapse.

Contributions of serotonin in addiction vulnerability.

The serotonin (5-hydroxytryptamine; 5-HT) system has long been associated with mood and its dysregulation implicated in the pathophysiology of mood and anxiety disorders. While modulation of 5-HT neurotransmission by drugs of abuse is also recognized, its role in drug addiction and vulnerability to drug relapse is a more recent focus of investigation. First, we review preclinical data supporting the serotonergic raphe nuclei and their forebrain projections as targets of drugs of abuse, with emphasis on the effects of psychostimulants, opioids and ethanol. Next, we examine the role of 5-HT receptors in impulsivity, a core behavior that contributes to the vulnerability to addiction and relapse. Finally, we discuss evidence for serotonergic dysregulation in comorbid mood and addictive disorders and suggest novel serotonergic targets for the treatment of addiction and the prevention of drug relapse.

Fractalkine/CX3CL1 enhances GABA synaptic activity at serotonin neurons in the rat dorsal raphe nucleus.

Serotonin (5-hydroxytryptamine; 5-HT) has an important role in mood regulation, and its dysfunction in the central nervous system (CNS) is associated with depression. Reports of mood and immune disorder co-morbidities indicate that immune-5-HT interactions may mediate depression present in immune compromised disease states including HIV/AIDS, multiple sclerosis, and Parkinson's disease. Chemokines, immune proteins that induce chemotaxis and cellular adhesion, and their G-protein coupled receptors distribute throughout the CNS, regulate neuronal patterning, and mediate neuropathology. The purpose of this study is to investigate the neuroanatomical and neurophysiological relationship between the chemokine fractalkine/CX3CL1 and its receptor CX3CR1 with 5-HT neurons in the rat midbrain raphe nuclei (RN). Immunohistochemistry was used to examine the colocalization of CX3CL1 or CX3CR1 with 5-HT in the RN, and whole-cell patch-clamp recordings in rat brain slices were used to determine the functional impact of CX3CL1 on 5-HT dorsal raphe nucleus (DRN) neurons. Greater than 70% of 5-HT neurons colocalize with CX3CL1 and CX3CR1 in the RN. CX3CL1 localizes as discrete puncta throughout the cytoplasm, whereas CX3CR1 concentrates to the perinuclear region of 5-HT neurons and exhibits microglial expression. CX3CL1 and CX3CR1 also colocalize with one another on individual RN cells. Electrophysiology studies indicate a CX3CL1-mediated enhancement of spontaneous inhibitory postsynaptic current (sIPSC) amplitude and dose-dependent increase of evoked IPSC (eIPSC) amplitude without affecting eIPSC paired-pulse ratio, a finding observed selectively in 5-HT neurons. CX3CL1's effect on eIPSC amplitude is blocked by pretreatment with an anti-CX3CL1 neutralizing antibody. Thus, CX3CL1 enhances postsynaptic GABA receptor number or sensitivity on 5-HT DRN neurons under conditions of both spontaneous and synaptically-evoked GABA release. CX3CL1 may indirectly inhibit 5-HT neurotransmission by increasing the sensitivity of 5-HT DRN neurons to GABA inputs. Therapies targeting CX3CL1 may treat serotonin related mood disorders, including depression experienced by patients with compromised immune systems.

Distinguishing characteristics of serotonin and non-serotonin-containing cells in the dorsal raphe nucleus: electrophysiological and immunohistochemical studies.

The membrane properties and receptor-mediated responses of rat dorsal raphe nucleus neurons were measured using intracellular recording techniques in a slice preparation. After each experiment, the recorded neuron was filled with neurobiotin and immunohistochemically identified as 5-hydroxytryptamine (5-HT)-immunopositive or 5-HT-immunonegative. The cellular characteristics of all recorded neurons conformed to previously determined classic properties of serotonergic dorsal raphe nucleus neurons: slow, rhythmic activity in spontaneously active cells, broad action potential and large afterhyperpolarization potential. Two electrophysiological characteristics were identified that distinguished 5-HT from non-5-HT-containing cells in this study. In 5-HT-immunopositive cells, the initial phase of the afterhyperpolarization potential was gradual (tau=7.3+/-1.9) and in 5-HT-immunonegative cells it was abrupt (tau=1.8+/-0.6). In addition, 5-HT-immunopositive cells had a shorter membrane time constant (tau=21.4+/-4.4) than 5-HT-immunonegative cells (tau=33.5+/-4.2). Interestingly, almost all recorded neurons were hyperpolarized in response to stimulation of the inhibitory 5-HT(1A) receptor. These results suggested that 5-HT(1A) receptors are present on non-5-HT as well as 5-HT neurons. This was confirmed by immunohistochemistry showing that although the majority of 5-HT-immunopositive cells in the dorsal raphe nucleus were double-labeled for 5-HT(1A) receptor-IR, a small but significant population of 5-HT-immunonegative cells expressed the 5-HT(1A) receptor. These results underscore the heterogeneous nature of the dorsal raphe nucleus and highlight two membrane properties that may better distinguish 5-HT from non-5-HT cells than those typically reported in the literature. In addition, these results present electrophysiological and anatomical evidence for the presence of 5-HT(1A) receptors on non-5-HT neurons in the dorsal raphe nucleus.

Effects of corticotropin-releasing factor on neuronal activity in the serotonergic dorsal raphe nucleus.

The present study examined the regional localization of corticotropin-releasing factor (CRF)- and 5-hydroxytryptamine (5-HT)-immunoreactive (IR) fibers within the rat dorsal raphe nucleus (DRN) using immunohistochemistry. Additionally, the effects of CRF, administered intracerebroventricularly (0.1-3.0 micrograms) or intraraphe (0.3-30 ng), on discharge rates of putative 5-HT DRN neurons were quantified using in vivo single unit recording in halothane-anesthetized rats. CRF-IR fibers were present at all rostrocaudal levels of the DRN and exhibited a topographical distribution. CRF produced predominantly inhibitory effects on DRN discharge at lower doses and these effects diminished or became excitatory at higher doses. Inhibition of DRN discharge by CRF was attenuated by the nonselective CRF antagonist, DPheCRF12-41 and the CRF-R1-selective antagonist, antalarmin, implicating the CRF-R1 receptor subtype in these electrophysiological effects. The present findings provide anatomical and physiological evidence for an impact of CRF on the DRN-5HT system.

Unrecognized high blood pressure. A major public health issue for the workplace.

Hypertension continues to be prevalent in the general population despite the public's increased awareness of cardiovascular disease. Population-wide detection and prevention of hypertension are high priority goals within preventive health care. According to recent National Heart, Lung, and Blood Institute (NHLBI) guidelines, high normal blood pressure (BP) (systolic 130 to 139 mm Hg or diastolic 85 to 89 mm Hg) is not an innocuous condition (NHLBI, 1997). High normal BP is a detectable, modifiable, antecedent condition to overt hypertension. Little is known about the incidence of high normal BP in the general population and of its relationship to stress. This study examined the prevalence of high normal and hypertensive levels of blood pressure in a convenience sample of 94 volunteer employees from a midsize corporation. Blood pressure and level of reported stress were assessed. Findings revealed rates of 11% and 30% high normal and hypertensive blood pressure levels, respectively. Ninety-six percent of participants assumed their blood pressures were normal. As in other studies, those employees with hypertensive blood pressure reported higher stress levels than normotensive employees. However, the population with high normal BP did not report significantly higher stress levels than normotensive employees. These findings suggest high normal and hypertensive blood pressures are prevalent cardiovascular disease risk factors among employees in the workplace. Most employees are unaware of their elevated BP and the risk of high normal BP. Occupational health nurses are in a strategic position to take a proactive approach to population-wide hypertension prevention by initiating worksite BP screening and education programs.

Serotonergic mediation of the effects of fluoxetine, but not desipramine, in the rat forced swimming test.

RATIONALE: The forced swimming test (FST) is a behavioral test in rodents that predicts the clinical efficacy of many types of antidepressant treatments. Recently, a behavior sampling technique was developed that scores individual response categories, including swimming, climbing and immobility. Although all antidepressant drugs reduce immobility in the FST, at least two distinct active behavioral patterns are produced by pharmacologically selective antidepressant drugs. Serotonin-selective reuptake inhibitors increase swimming behavior, while drugs acting primarily to increase extracellular levels of norepinephrine or dopamine increase climbing behavior. Distinct patterns of active behaviors in the FST may be mediated by distinct neurotransmitters, but this has not been shown directly. OBJECTIVES: The present study examined the role of serotonin in mediating active behaviors in the forced swimming test after treatment with two antidepressant drugs, the selective serotonin reuptake inhibitor, fluoxetine and the selective norepinephrine reuptake inhibitor, desipramine. METHODS: Endogenous serotonin was depleted by administering para-cholorophenylalanine (PCPA, 150 mg/kg, IP.) to rats 72 h and 48 h prior to the swim test. Fluoxetine (10 mg/kg, SC) or desipramine (10 mg/kg, SC) was given three times over a 24-h period prior to the FST. Behavioral responses, including immobility, swimming and climbing, were counted during the 5-min test. RESULTS: Pretreatment with PCPA blocked fluoxetine-induced reduction in immobility and increase in swimming behavior during the FST. In contrast, PCPA pretreatment did not interfere with the ability of desipramine to reduce immobility and increase climbing behavior. CONCLUSIONS: Depletion of serotonin prevented the behavioral effects of the selective serotonin reuptake inhibitor fluoxetine in the rat FST. Furthermore, depletion of serotonin had no impact on the behavioral effects induced by the selective norepinephrine reuptake inhibitor, desipramine. The effects of antidepressant drugs on FST-induced immobility may be exerted by distinguishable contributions from different neurotransmitter systems.

Effects of corticotropin-releasing factor on brain serotonergic activity.

The serotonergic dorsal raphe nucleus is innervated by corticotropin-releasing factor (CRF) and expresses CRF receptors, suggesting that endogenous CRF impacts on this system. The present study characterized interactions between CRF and the dorsal raphe serotonin (5-HT) system. The effects of intracerebroventricularly (i.c.v.) administered CRF on microdialysate concentrations of 5-HT in the lateral striatum of freely moving rats were determined. CRF had biphasic effects, with 0.1 and 0.3 microgram decreasing, and 3.0 micrograms increasing 5-HT dialysate concentrations. i.c.v. administration of CRF inhibited neuronal activity of the majority of dorsal raphe neurons at both low (0.3 microgram) and high (3 micrograms) doses. Likewise, intraraphe administration of CRF (0.3 and 1.0 ng) had predominantly inhibitory effects on discharge rate. Together, these results suggest that CRF is positioned to regulate the function of the dorsal raphe serotonergic system via actions within the cell body region. This regulation may play a role in stress-related psychiatric disorders in which 5-HT has been implicated.

The effect of repeated exposure to forced swimming on extracellular levels of 5-hydroxytryptamine in the rat.

The effects of repeated exposure to forced swimming was examined on extracellular concentrations of 5-hydroxytryptamine (5-HT), behavioral, and endocrine responses in rats. Animals were exposed to a 15-min swimming session on two consecutive days. On the first day, the swimming session increased extracellular 5-HT by 80 % over baseline in the striatum and reduced 5-HT to 40 % below baseline in the lateral septum. On the second day, however, the swimming session produced no effect on 5-HT in either brain region. Ratings of behavior showed that rats demonstrated climbing and swimming behaviors and developed immobility during the initial swimming session and that ratings of immobility increased and swimming decreased during the second swimming session. Immobility was positively correlated and swimming was negatively correlated with changes in extracellular 5-HT in the lateral septum but not in the striatum. Plasma corticosterone was equally elevated by 950 % after either 1 or 2 days of swimming exposure. These results show that there is rapid adaptation to the effects of repeated forced swimming on the regionally-specific, bi-directional response of extracellular 5-HT. In addition, changes in extracellular 5-HT in the lateral septum may be related to the behaviors produced during the forced swimming test that underlie its utility as an animal model of depression.

Interaction between the forced swimming test and fluoxetine treatment on extracellular 5-hydroxytryptamine and 5-hydroxyindoleacetic acid in the rat.

We used in vivo microdialysis to examine extracellular levels of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) in the striatum and the lateral septum during the forced swimming test, (FST) a behavioral test conducted in rats that is commonly used to predict the effect of antidepressant drugs. The forced swimming test consisted of a 15-min pretest swim and a 5-min test swim 24 hr later. The antidepressant fluoxetine (20 mg/kg s.c.) or saline was administered 23.5, 5 and 1 hr before the test swim. In the striatum, the pretest swim increased 5-HT in both treatment groups. On the second day, the test swim had no effect on 5-HT in saline-treated rats but slightly decreased striatal 5-HT in fluoxetine-treated rats. In the lateral septum, the pretest swim decreased 5-HT in both treatment groups. On the second day, the test swim had no effect on 5-HT in saline-treated rats but decreased lateral septum 5-HT in fluoxetine-treated rats. Ratings of behavior showed that fluoxetine treatment increased swimming behavior and decreased immobility during the test swim. Immobility was positively correlated and swimming was negatively correlated with changes in extracellular 5-HT in the lateral septum but not in the striatum. Therefore, fluoxetine treatment altered adaptation of the regional response of extracellular 5-HT ordinarily produced in the FST, reversing the 5-HT response to the initial swim in the striatum and restoring the response to the initial swim in the lateral septum.

The effects of different stressors on extracellular 5-hydroxytryptamine and 5-hydroxyindoleacetic acid.

The effects of application of five different stressors on extracellular 5-hydroxytryptamine and 5-hydroxyindoleacetic acid in the striatum and hippocampus were compared using in vivo microdialysis. Forced swimming for 30 min elevated extracellular 5-hydroxytryptamine to 90% above basal levels and reduced 5-hydroxyindoleacetic acid to 45% of basal levels in the striatum during the swim session. In contrast, hippocampal 5-hydroxytryptamine was not altered significantly by forced swimming but 5-hydroxyindoleacetic acid levels were reduced to 60% of basal levels. Tail pinch for 5 min elevated 5-hydroxytryptamine to 55% above basal levels in striatum and to 35% above basal levels in hippocampus. Tail pinch had no effect on 5-hydroxyindoleacetic acid in either brain region. In contrast to forced swimming and the tail pinch, the other three stressors, immobilization stress for 100 min, exposure to a cold environment (4 degrees C) for 2 h, and forced motor activity on a rotarod for 30 min, failed to alter extracellular 5-hydroxytryptamine in either the striatum or the hippocampus. All five stressors increased plasma corticosterone levels: tail pinch, 246%; cold stress, 432%; immobilization, 870%; forced motor activity, 1030%; and forced swimming, 1530%. These results suggest that individual stressors produce different effects on extracellular 5-hydroxytryptamine in different brain regions. In addition, there does not appear to be a relationship between the effects of stressors on the 5-hydroxytryptamine system and the magnitude of their ability to activate the hypothalamic-pituitary-adrenal axis.

Effect of destruction of serotonin neurons on basal and fenfluramine-induced serotonin release in striatum.

This study examined the relationship between the magnitude of tissue serotonin (5-HT) depletion produced by treatment with the neurotoxin 5,7-dihydroxytryptamine (5,7-DHT) and basal and fenfluramine-induced 5-HT release in the striatum. Separate groups of rats were treated with either vehicle or 5,7-DHT (100 micrograms: 76% striatal 5-HT depletion; or 200 micrograms: 93% striatal 5-HT depletion). Four weeks after treatment, 5-HT release was measured in the ventral striatum using in vivo microdialysis in animals anesthetized with chloral hydrate. Basal 5-HT levels were not significantly altered in any lesion group, whereas basal 5-hydroxyindoleacetic acid levels were dose-dependently reduced by 5,7-DHT. In contrast, the increase of 5-HT release produced by fenfluramine treatment (10 mg/kg) was diminished significantly after 5-HT neuronal destruction in correlation with the reduction of striatal tissue 5-HT content. Fractional 5-HT efflux, a measure of the 5-HT release from surviving striatal nerve terminals, was also significantly elevated when tissue depletion of 5-HT exceeded 95%. This study suggests that compensatory mechanisms may enable surviving 5-HT terminals to maintain basal 5-HT levels in the striatum with as little as 5% of the terminals remaining, but those mechanisms are not sufficient to allow the damaged system to respond to a pharmacological challenge.

Regional differences in the effects of forced swimming on extracellular levels of 5-hydroxytryptamine and 5-hydroxyindoleacetic acid.

The effects of forced swimming for 30 min on extracellular 5-hydroxytryptamine (5-HT) levels were examined in five brain regions in rats using in vivo microdialysis. A single dialysis probe was implanted under surgical anesthesia into either the striatum, ventral hippocampus, frontal cortex, amygdala, or lateral septum on the day before the study. Dialysate content of 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) was measured by HPLC. Forced swimming elevated extracellular levels of 5-HT in the striatum to a maximum of 90% above baseline. In contrast, forced swimming reduced 5-HT levels in the amygdala and lateral septum to 50 and 40% of baseline, respectively. In the hippocampus and frontal cortex, 5-HT levels were not altered significantly by forced swimming. In all five brain regions, forced swimming reduced 5-HIAA levels to 45-60% of baseline. These results suggest that forced swimming modulates 5-HT neurotransmission in a regionally specific manner. Aside from being a significant biological stressor, the forced swimming test is used as an animal behavioral model to detect antidepressant drugs, including drugs that alter 5-HT neurotransmission. It is possible that the alterations of extracellular levels of 5-HT produced by forced swimming in certain brain regions may be associated with the ability of antidepressant drugs to selectively alter behavioral performance during the forced swimming test.

Discriminative stimulus properties of the benzodiazepine receptor inverse agonist methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM).

The purpose of this study was to determine whether rats could be trained to discriminate the stimulus properties of the benzodiazepine (BZ) receptor inverse agonist DMCM from saline in a conditioned taste aversion paradigm. On a drug trial, water-deprived rats were injected with DMCM (0.55-0.6 mg/kg IP), allowed access to a 0.25% saccharin solution for 30 min, and then injected with LiCl. On non-drug trials, saline injections bracketed the drinking period. Conditioned controls were treated similarly with DMCM and saline on drug and non-drug trials, but received injections of saline instead of LiCl. At the completion of training, CMCM produced a 69% reduction of saccharin consumption on drug trials, compared with 23% for conditioned controls. The stimulus properties of DMCM were then measured by its ability to reduce the preference for saccharin over water in a two-bottle choice test. DMCM reduced saccharin preference in rats that received discrimination training from 68% to 19%, but did not alter saccharin preference in conditioned controls. Other compounds with varying activity at BZ receptors were evaluated for their ability to substitute for the discriminative stimulus effects of DMCM. Two BZ receptor inverse agonists, beta-CCE (10-18 mg/kg) and FG 7142 (3.2-18 mg/kg), substituted completely for DMCM. Partial substitution for DMCM was shown by the BZ receptor antagonist CGS 8216 (3.2-10 mg/kg) and the non-BZ convulsant pentylenetetrazol (10-20 mg/kg). The BZ receptor agonists chlordiazepoxide (0.32-5.0 mg/kg), diazepam (0.32-10 mg/kg), and alprazolam (0.1-3.2 mg/kg) and the BZ receptor antagonist flumazenil (1.0-32 mg/kg) failed to substitute for the DMCM stimulus. Pretreatment with flumazenil (1.0 mg/kg) blocked the stimulus effects of the training dose of DMCM and produced a shift to the right of the DMCM generalization curve. The pattern of compounds that substituted for the DMCM stimulus and the blockade of that stimulus by flumazenil indicate that the stimulus properties of DMCM are associated with its effects as a BZ receptor inverse agonist.