Taleen Hanania

Translational aspects of pharmacological research into anxiety disorders: The stress-induced hyperthermia (SIH) paradigm

In anxiety research, the search for models with sufficient clinical predictive validity to support the translation of animal studies on anxiolytic drugs to clinical research is often challenging. This review describes the stress-induced hyperthermia (SIH) paradigm, a model that studies the activation of the autonomic nervous system in response to stress by measuring body temperature. The reproducible and robust SIH response, combined with ease of testing, make the SIH paradigm very suitable for drug screening. We will review the current knowledge on the neurobiology of the SIH response, discuss the role of GABA(A) and serotonin (5-HT) pharmacology, as well as how the SIH response relates to infectious fever. Furthermore, we will present novel data on the SIH response variance across different mice and their sensitivity to anxiolytic drugs. The SIH response is an autonomic stress response that can be successfully studied at the level of its physiology, pharmacology, neurobiology and genetics and possesses excellent animal-to-human translational properties.

SAHA Enhances Synaptic Function and Plasticity In Vitro but Has Limited Brain Availability In Vivo and Does Not Impact Cognition

Suberoylanilide hydroxamic acid (SAHA) is an inhibitor of histone deacetylases (HDACs) used for the treatment of cutaneous T cell lymphoma (CTCL) and under consideration for other indications. In vivo studies suggest reducing HDAC function can enhance synaptic function and memory, raising the possibility that SAHA treatment could have neurological benefits. We first examined the impacts of SAHA on synaptic function in vitro using rat organotypic hippocampal brain slices. Following several days of SAHA treatment, basal excitatory but not inhibitory synaptic function was enhanced. Presynaptic release probability and intrinsic neuronal excitability were unaffected suggesting SAHA treatment selectively enhanced postsynaptic excitatory function. In addition, long-term potentiation (LTP) of excitatory synapses was augmented, while long-term depression (LTD) was impaired in SAHA treated slices. Despite the in vitro synaptic enhancements, in vivo SAHA treatment did not rescue memory deficits in the Tg2576 mouse model of Alzheimer’s disease (AD). Along with the lack of behavioral impact, pharmacokinetic analysis indicated poor brain availability of SAHA. Broader assessment of in vivo SAHA treatment using high-content phenotypic characterization of C57Bl6 mice failed to demonstrate significant behavioral effects of up to 150 mg/kg SAHA following either acute or chronic injections. Potentially explaining the low brain exposure and lack of behavioral impacts, SAHA was found to be a substrate of the blood brain barrier (BBB) efflux transporters Pgp and Bcrp1. Thus while our in vitro data show that HDAC inhibition can enhance excitatory synaptic strength and potentiation, our in vivo data suggests limited brain availability may contribute to the lack of behavioral impact of SAHA following peripheral delivery. These results do not predict CNS effects of SAHA during clinical use and also emphasize the importance of analyzing brain drug levels when interpreting preclinical behavioral pharmacology.

The triple monoaminergic reuptake inhibitor DOV 216,303 has antidepressant effects in the rat olfactory bulbectomy model and lacks sexual side effects

Current antidepressants have a delayed onset of action and disturbing side effects, including inhibition of sexual behavior. It is hypothesized that novel drugs, hitting multiple disease-relevant targets, may yield a new generation of superior antidepressants. One such approach is simultaneous inhibition of serotonin, norepinephrine and dopamine transporters. We tested the triple uptake inhibitor (TUI), DOV 216,303 (5, 10 and 20 mg/kg) after 1, 7 and 14 days administration in the olfactory bulbectomized (OBX) rat depression model, and in a model of rat sexual behavior to detect putative sexual side effects. Chronic, but not acute treatment of DOV 216,303 (20 mg/kg) normalized OBX-induced hyperactivity in the open field, similar to the effect of imipramine (20 mg/kg). None of the doses of DOV 216,303 had any effect on sexual behavior at any time point. The results indicate that DOV 216,303 displays antidepressant efficacy and is devoid of sexual side effects.

Sub-optimal performance in the 5-choice serial reaction time task in rats was sensitive to methylphenidate, atomoxetine and D-amphetamine, but unaffected by the COMT inhibitor tolcapone

Prefrontal cortical dopamine (DA) and norepinephrine (NE) are implicated in multiple aspects of cognitive function assessed via the 5-choice serial reaction time task (5-CSRTT) in rodents. The present studies assessed the effects of the NE reuptake inhibitor atomoxetine (0.5-2.0 mg/kg), the mixed DA/NE reuptake inhibitor methylphenidate (0.1-2.0 mg/kg), the catecholamine releaser D-amphetamine (0.1-1.0 mg/kg) and the catecholamine-o-methyl-transferase (COMT) inhibitor tolcapone (3.0-30.0 mg/kg) in rats that exhibited sub-optimal performance (reduced accuracy: <70% correct) in the 5-CSRTT. Increased ITI durations were associated with increased premature responding. Decreased ITI durations resulted in increased percent omissions, increased perseverative responses and increased response latencies, but had no effects on magazine latencies or percent correct. Atomoxetine decreased premature responding at prolonged ITI durations and methylphenidate decreased percent omissions at low doses (0.1 and 0.5 mg/kg). By contrast, D-amphetamine increased premature and perseverative responding in a dose-dependent manner (0.3-1.0 mg/kg). Finally, tolcapone had no effects on sub-optimal performance in the variable ITI 5-CSRTT. These results suggest minimal potential of tolcapone as a therapeutic agent for ADHD and implicate cortical NE, not DA, in impulsive action.

Psychostimulant-like discriminative stimulus and locomotor sensitization properties of the wake-promoting agent modafinil in rodents

UNLABELLED The present studies assessed the potential abuse liability and likely mechanism(s) of action of the wake-promoting agent modafinil. METHODS Experiments assessed the locomotor sensitization (LS) and discriminative stimulus (DS) properties of modafinil in mouse and rat, respectively. Comparative data were generated with a range of psychostimulants and monoamine reuptake inhibitors. RESULTS Repeated administration of d-amphetamine and cocaine, psychostimulants with high abuse liability, resulted in the induction and expression of LS in mice. Bupropion and caffeine, two psychostimulants not abused in humans, were not associated with LS. GBR12909 induced LS during repeated exposure, but there was no evidence of expression of LS after acute challenge following withdrawal. In contrast, repeated administration of modafinil resulted in the expression, but not induction, of LS. d-amphetamine, but not the mu-opioid agonist morphine or the nAChR agonist nicotine, fully substituted for the cocaine DS in rats. The selective dopamine transporter (DAT) inhibitor GBR12909 fully substituted, the preferential norepinephrine transporter (NET) inhibitor desipramine partially substituted, and the selective serotonin reuptake inhibitor citalopram failed to substitute for cocaine. Modafinil fully substituted for cocaine, similar to the mixed DAT/NET inhibitor bupropion. CONCLUSIONS Two preclinical assays indicated potential abuse liability of modafinil; drug discrimination studies suggest DAT blockade by modafinil is a likely mechanism of action in vivo.

Dissociation between duration of action in the forced swim test in mice and nicotinic acetylcholine receptor occupancy with sazetidine, varenicline, and 5-I-A85380

RationaleNicotinic acetylcholine receptor (nAChR) agonists, partial agonists, and antagonists have antidepressant-like effects in rodents and reduce symptoms of depression in humans.ObjectivesThe study determined whether the antidepressant-like effect of the nAChR β2* partial agonist sazetidine-A (sazetidine) in the forced swim test was due to activation or desensitization of β2* nAChRs. The study also determined if sazetidine’s behavioral responses in the forced swim test corresponded to β2* nAChRs receptor occupancy and drug bioavailability.ResultsAcute antidepressant-like effects in the forced swim test were seen with sazetidine and the full β2* agonist 5-I-A8350 (BALB/cJ mice) and the less selective β2* partial agonist varenicline in C57BL/6J but not BALB/cJ mice. The role of β2* nAChRs was confirmed by results showing: (1) reversal of sazetidine’s antidepressant-like effects in the forced swim test by nAChR antagonists mecamylamine and dihydro-β-erythroidine; (2) absence of sazetidine’s effect in mice lacking the β2 subunit of the nAChR; and (3) a high correspondence between behaviorally active doses of sazetidine and β2* receptor occupancy. β2* receptor occupancy following acute sazetidine, varenicline, and 5-I-A8350 lasted beyond the duration of action in the forced swim test. Sazetidine’s long lasting receptor occupancy did not diminish behavioral efficacy in the forced swim test following repeated dosing.ConclusionsResults demonstrate that activation of a small population of β2* nAChRs (10–40%) is sufficient to elicit sazetidine’s antidepressant-like actions without producing tolerance and suggest that ligands that activate β2* nAChRs would be promising targets for the development of a new class of antidepressant.

Augmentation of locomotor activity by chronic phencyclidine is associated with an increase in striatal NMDA receptor function and an upregulation of the NR1 receptor subunit

Phencyclidine (PCP) is a drug of abuse that produces schizophrenia‐like symptoms in humans and increases locomotor activity and stereotypic behavior in rodents. PCP‐induced alteration in rat locomotor activity is thought to be mediated by an inhibition of N‐methyl‐D‐aspartate (NMDA) receptors in the striatum and other brain regions. In this study, rats treated chronically with PCP (20 mg/kg once per day for 5 days) showed a marked increase in locomotor activity following a PCP challenge (3.2 mg/kg) administered after either 3 or 8 days of withdrawal. In biochemical assays, the release of striatal [14C]GABA by NMDA was enhanced by about 77% by chronic PCP treatment, whereas [3H]ACh release was increased by about 31% in tissue from PCP‐treated rats. Even though binding experiments with 1‐[1‐(2‐thiehyl)cyclohexyl]piperidyl‐3,4 3H(N) ([3H]TCP) showed no alteration in the Kd or Bmax in whole striatum, quantitative immunocytochemical experiments found an upregulation in the NR1 subunit in the cell bodies and neuropil of cortical and striatal regions of the forebrain following chronic PCP treatment. An increase in the size of NR1‐immunoreactive cells in the forebrain was also observed following chronic PCP treatment. Together, these data may help in understanding the mechanisms underlying the adaptive response to chronic reduction in glutamatergic NMDA transmission that has been postulated to be involved in the etiology of schizophrenia. Synapse 31:229–239, 1999.

Impulsive action and impulsive choice are mediated by distinct neuropharmacological substrates in rat.

Impulsivity is a heterogeneous construct according to clinical and preclinical behavioural measures and there is some preliminary evidence indicating distinct neurobiological substrates underlying the sub-components of impulsivity. Two preclinical assays, the five-choice serial reaction time task (5-CSRTT) and the delayed discounting task (DDT), are hypothesized to provide measures of impulsive action (premature responding) and impulsive choice (percent choice for delayed reward), respectively. In the present studies, we show that the norepinephrine reuptake inhibitor atomoxetine attenuated premature responding in the 5-CSRTT, but was ineffective in the DDT. The mixed dopamine/norepinephrine reuptake inhibitor methylphenidate exhibited an opposite profile of effects. In addition, blockade of 5-HT2A/C receptors via ketanserin decreased premature responding but had no effects on percent choice for delayed reward; blockade of 5-HT2C receptors via SB 242084 had opposite effects. Follow-up studies provided some limited evidence of additive effects of 5-HT2A/C receptor blockade on the effects of atomoxetine on impulsive action. These studies demonstrate dissociable profiles of stimulant vs. non-stimulant attention deficit hyperactivity disorder medications and 5-HT subtype-selective ligands, in the 5-CSRTT and DDT assays. Thus, the present findings support the sub-categorization of impulsivity and suggest that 5-HT receptor subtype-selective antagonists may provide therapeutic targets for disorders characterized by different forms of impulsivity.

Pharmacological characterization of harmaline-induced tremor activity in mice.

Harmaline-induced tremor in rodents is a model of essential tremor. We utilized a novel assay to quantify tremor activity in mice and found that tremor activity was dependent on harmaline dose. The first-line clinical essential tremor treatments propranolol, primidone and gabapentin and gamma-hydroxybutyrate (GHB) significantly attenuated harmaline-induced tremor. The anticonvulsants valproate and carbamazepine and the mood stabilizer lithium suppressed harmaline-induced tremor. The gamma-amino-butyric acid (GABA) receptor subtype A receptor agonist muscimol attenuated harmaline-induced tremor. By contrast, the GABA(B) receptor agonist R-baclofen increased tremor at the lowest dose tested, but had no effects at higher doses. Administration of the non-competitive N-methyl-D-aspartate (NMDA) receptor antagonists phencyclidine or 5R,10S-(+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine hydrogen maleate (MK-801) attenuated harmaline-induced tremor. The competitive NMDA antagonist D-4-[(2E)-3-phosphono-2-propenyl]-2-piperazinecarboxylic acid (d-CPPene) dose-dependently blocked harmaline-induced tremor, as did the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide disodium salt (NBQX). The metabotropic glutamate subtype 5 (mGlu5) receptor antagonist 6-methyl-2-(phenylethynyl)pyridine (MPEP) was inactive against tremor. The dopamine reuptake inhibitor GBR12909 and the dopamine D(1)/D(2) receptor agonist apomorphine attenuated harmaline-induced tremor. Follow-up studies indicated that dopamine D(2)/D(3) but not dopamine D(1) receptor activation likely mediates the effects of apomorphine and GBR12909. Administration of compounds with sedative side-effects had no effect on tremor activity. In summary, the present data confirm the pharmacological validity of harmaline-induced tremor in mice, quantified via a novel assay, as an animal model of essential tremor. Further, these data provide additional evidence for the roles of ionotropic glutamate, GABA(A) and dopamine D(2)/D(3) receptors in the neurobiology of harmaline-induced tremor.

Regulation of NMDA-stimulated [14C]GABA and [3H]acetylcholine release by striatal glutamate and dopamine receptors

Striatal function is heavily influenced by glutamatergic and dopaminergic afferent input. To ultimately better understand how the N-methyl-D-aspartate (NMDA) antagonist, phencyclidine (PCP), alters striatal function, we sought to determine how NMDA receptor function is influenced by activation of other glutamatergic receptors and by dopaminergic receptors. To this end, we used NMDA-stimulated efflux of [14C]GABA and [3H]acetylcholine (ACh) from striatal slices to assess the influence of these receptors on NMDA function. NMDA-stimulated [14C]GABA release was more sensitive to NMDA and glycine antagonists than was [3H]ACh release, suggesting that different NMDA receptors regulate the release of these neurotransmitters. Furthermore, NMDA-stimulated [3H]ACh release was inhibited by a D2 receptor mechanism whereas NMDA-stimulated [14C]GABA release was enhanced by D1 receptor activation. NMDA and (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid hydrobromide (AMPA) interact additively to evoke [3H]ACh release, and synergistically to evoke [14C]GABA release. An additive effect of NMDA and kainate (KA) was found on [14C]GABA release, but NMDA and KA acted in a less than additive manner in evoking [3H]ACh release. KA-stimulated [3H]ACh release was largely blocked by NMDA antagonists, suggesting mediation through activation of NMDA receptors, probably secondary to KA-induced glutamate release. A selective group II metabotropic receptor agonist inhibited NMDA-stimulated [14C]GABA and [3H]ACh release. On the other hand, NMDA-stimulated [14C]GABA release was potentiated by activation of group I metabotropic receptors. Thus, in addition to the differential modulation by D1- and D2-like receptors, the release of striatal neurotransmitters by NMDA receptor activation depends on the extent to which the other glutamate receptors, both ionotropic and metabotropic, are activated.