Modulation of the human ρ1 GABAA receptor by inhibitory steroids

Rationale

Modulators of the ρ1 GABA_A receptor may be useful in the treatment of visual, sleep, and cognitive disorders. Neuroactive steroids and analogues have been shown to modulate ρ1 receptor function, but the molecular mechanisms are poorly understood.

Objectives

We employed electrophysiology and voltage-clamp fluorometry to compare the actions of several neuroactive steroids and analogues on the human ρ1 GABA_A receptor.

Results

Results confirmed that P294S and T298F mutations affect modulation by steroids. The P294S mutation abolished inhibition by (3α,5β)-3-hydroxypregnan-20-one (3α5βP) while the T298F mutation eliminated inhibition by 17β-estradiol. Voltage-clamp fluorometry demonstrated that steroids differing in the presence of a charged group on C3 or nature of substituent on C17 uniquely modified fluorescence changes elicited by GABA in the extracellular domain. The I307Q mutation reversed the inhibitory effect of 3α5βP but was without effect on modulation by (3α,5β)-3-hydroxypregnan-20-one sulfate or 17β-estradiol. The effect of 3α5βP on the fluorescence change generated at Y241C was dependent on whether the steroid acted as an inhibitor or a potentiator. Further, the effect was limited to uncharged 5β-reduced steroids containing an acetyl group on C17.

Conclusions

The data demonstrate that steroids and analogues differ with respect to conformational changes elicited by these drugs as well as sensitivity to the effects of mutations. Steroids and analogues could be provisionally divided into three major groups based on their actions on the ρ1 GABA_A receptor: 5β-reduced uncharged steroids, sulfated and carboxylated steroids, and 17β-estradiol. Further division among 5β-reduced uncharged steroids was based on substituent at position C17.     

Effects of inhibitory GABA-active neurosteroids on cocaine seeking and cocaine taking in rats

Rationale

Several compounds that potentiate GABA-induced inhibitory currents also decrease stress, anxiety and addiction-related behaviors. Because of the well-established connection between stress and addiction, compounds that reduce stress-induced responses might be efficacious in treating addiction. Since endogenous neurosteroids such as allopregnanolone may function in a manner similar to benzodiazepines to reduce HPA axis activation and anxiety following stressful stimuli, we hypothesized that exogenously applied neurosteroids would reduce cocaine reinforcement in two animal models.

Methods

Male Wistar rats were trained to self-administer cocaine and food under a concurrent alternating operant schedule of reinforcement. Two separate groups of rats were trained to self-administer cocaine or food pellets and were then exposed to similar cue-induced reinstatement paradigms. Both groups of rats were pretreated with various doses of neurosteroids.

Results

Allopregnanolone and 3α-hydroxy-3β-methyl-17β-nitro-5α-androstane (R6305-7, a synthetic neurosteroid) were ineffective in selectively decreasing cocaine relative to food self-administration. On the other hand, both allopregnanolone and R6305-7 significantly decreased the cue-induced reinstatement of extinguished cocaine seeking, confirmed by one-way ANOVA.

Conclusions

These results suggest that neurosteroids may be effective in reducing the relapse to cocaine use without affecting ongoing cocaine self-administration.     

11-trifluoromethyl-phenyldiazirinyl neurosteroid analogues: potent general anesthetics and photolabeling reagents for GABAA receptors

Rationale

While neurosteroids are well-described positive allosteric modulators of gamma-aminobutyric acid type A (GABA_A) receptors, the binding sites that mediate these actions have not been definitively identified.

Objectives

This study was conducted to synthesize neurosteroid analogue photolabeling reagents that closely mimic the biological effects of endogenous neurosteroids and have photochemical properties that will facilitate their use as tools for identifying the binding sites for neurosteroids on GABA_A receptors.

Results

Two neurosteroid analogues containing a trifluromethyl-phenyldiazirine group linked to the steroid C11 position were synthesized. These reagents, CW12 and CW14, are analogues of allopregnanolone (5α-reduced steroid) and pregnanolone (5β-reduced steroid), respectively. Both reagents were shown to have favorable photochemical properties with efficient insertion into the C–H bonds of cyclohexane. They also effectively replicated the actions of allopregnanolone and pregnanolone on GABA_A receptor functions: they potentiated GABA-induced currents in Xenopus laevis oocytes transfected with α₁β₂γ_2L subunits, modulated [³⁵S]t-butylbicyclophosphorothionate binding in rat brain membranes, and were effective anesthetics in Xenopus tadpoles. Studies using [³H]CW12 and [³H]CW14 showed that these reagents covalently label GABA_A receptors in both rat brain membranes and in a transformed human embryonal kidney (TSA) cells expressing either α₁ and β₂ subunits or β₃ subunits of the GABA_A receptor. Photolabeling of rat brain GABA_A receptors was shown to be both concentration-dependent and stereospecific.

Conclusions

CW12 and CW14 have the appropriate photochemical and pharmacological properties for use as photolabeling reagents to identify specific neurosteroid-binding sites on GABA_A receptors.     

The antidepressant-like effects of topiramate alone or combined with 17β-estradiol in ovariectomized Wistar rats submitted to the forced swimming test

Rationale

There is a significant delay in the clinical response of antidepressant drugs, and antidepressant treatments produce side effects.

Objective

We examined the relationship between 17β-estradiol and topiramate in ovariectomized Wistar rats submitted to the forced swimming test (FST).

Methods

Topiramate was administered alone or combined with 17β-estradiol to ovariectomized rats submitted to the FST.

Results

Topiramate (20 mg/kg, P?P?P?P?P?Conclusions Topiramate alone or combined with 17β-estradiol produced antidepressant-like actions; and 17β-estradiol shortened the onset of the antidepressant-like effects of topiramate.     

Characterization and Evaluation of Multi-Component Crystals of Hydrochlorothiazide

Purpose

The present work aims at improving the physicochemical properties of hydrochlorothiazide, a poorly water soluble antihypertensive drug by preparing its multi-component crystals with nicotinic acid (HCT-NA) and 2-picolinic acid (HCT-PIC).

Methods

The crystals prepared by solution crystallization were investigated by thermoanalytical techniques. The crystal structures of HCT-NA (1) and HCT-PIC (2) were determined by the single crystal X-ray diffraction and were assessed for their aqueous solubility, antihypertensive activity and acute toxicity in rats.

Results

Both 1 and 2 crystallized in the orthorhombic space group P2₁2₁2₁ and formation of salts were confirmed. The solubility profiles of 1 and 2 in basic media showed a maximum release of 2.5 mg/ml and 1.9 mg/ml, respectively, in comparison to the drug (0.82 mg/ml). The in-vivo antihypertensive activity of 1 in deoxycorticosterone acetate salt induced hypertensive rats showed 1.5 fold improvement. No increase in the signs of toxicity were revealed in rats during the acute toxicity studies even at doses of 2,000 mg/kg by body weight in comparison to the free drug. Histopathological findings supported the safety of these multi-component crystals.

Conclusions

The new solid phases exhibit potential to be explored for the oral drug delivery of HCT with improved solubility and therapeutic outcome.     

Pharmacological profile of a 17β-heteroaryl-substituted neuroactive steroid

Rationale

In order to improve upon the pharmacological properties of the neuroactive steroid ganaxolone, it was used as the starting point in the design of novel neurosteroids that replace the 17β-acetyl side chain with an isoxazole bioisostere.

Objectives

UCI-50027 (3-[3α-hydroxy-3β-methyl-5α-androstan-17β-yl]-5-(hydroxymethyl)isoxazole) was designed as an orally active neuroactive steroid specifically targeted at the gamma-aminobutyric acid(A) receptor (GABA_AR).

Methods

UCI-50027 was tested in vitro in Xenopus oocytes expressing human GABA_ARs and in vivo as an anticonvulsant, for ataxic effects and for anxiolytic activity.

Results

In vitro, UCI-50027 dose-dependently enhanced the activity of GABA at human α₁β₂γ_2L, α₂β₁γ_2L, and α₄β₃δ GABA_ARs. Consistent with its action as a positive allosteric modulator (PAM), it had no direct activity in the absence of GABA. UCI-50027 protected against acute pentylenetetrazol (PTZ)-induced convulsions with an ED₅₀ of 6 mg/kg p.o. In the rotarod (RR) paradigm in mice, the AD₅₀ (the ataxic dose where half of the animals fail the RR test) was found to be 38 mg/kg p.o., giving a therapeutic index (TI = RR AD₅₀/PTZ ED₅₀)～6 versus 2.8 for ganaxolone. In the mouse-elevated plus maze (EPM) model for anxiety, UCI-50027 showed a minimum effective dose (MED) ≤0.3 mg/kg p.o. Thus, the TI (TI = RR AD₅₀/EPM MED) for the compound as an anxiolytic is ≥127 versus 3.3 for ganaxolone.

Conclusions

UCI-50027 is an orally active neuroactive steroid with pharmacological activity consistent with a GABA_AR PAM that has an improved separation between anticonvulsant/anxiolytic and rotarod effects, potent activity as an anticonvulsant and anxiolytic when compared to ganaxolone.     

Anticonvulsant potencies of the enantiomers of the neurosteroids androsterone and etiocholanolone exceed those of the natural forms

Rationale

Androsterone [(3α,5α)-3-hydroxyandrostan-17-one; 5α,3α-A] and its 5β-epimer etiocholanolone [(3α,5β)-3-hydroxyandrostan-17-one; 5β,3α-A)], the major excreted metabolites of testosterone, are neurosteroid positive modulators of GABA_A receptors. Such neurosteroids typically show enantioselectivity in which the natural form is more potent than the corresponding unnatural enantiomer. For 5α,3α-A and 5β,3α-A, the unnatural enantiomers are more potent at GABA_A receptors than the natural forms.

Objectives

The aim of this study was to compare the anticonvulsant potencies and time courses of 5α,3α-A and 5β,3α-A with their enantiomers in mouse seizure models.

Methods

Steroids were administered intraperitoneally to male NIH Swiss mice 15 min (or up to 6 h in time course experiments) prior to administration of an electrical stimulus in the 6-Hz or maximal electroshock (MES) seizure tests or the convulsant pentylenetetrazol (PTZ).

Results

In the 6-Hz test, the ED₅₀ values of ent-5α,3α-A was 5.0 mg/kg whereas the value for 5α,3α-A was 12.1 mg/kg; the corresponding values in the PTZ seizure test were 22.8 and 51.8 mg/kg. Neurosteroid GABA_A receptor-positive allosteric modulators are generally weak in the MES seizure test and this was confirmed in the present study. However, the atypical relative potency relationship was maintained with ED₅₀ values of 140 and 223 mg/kg for ent-5α,3α-A and 5α,3α-A, respectively. Similar relationships were obtained for the 5β-isomers, except that the enantioselectivity was accentuated. In the 6-Hz and PTZ tests, the ED₅₀ values of ent-5β,3α-A were 11.8 and 20.4 mg/kg whereas the values for 5β,3α-A were 57.6 and 109.1 mg/kg. Protective activity in the 6-Hz test of ent-5α,3α-A persisted for somewhat longer (~5 h) than for 5α,3α-A (~4 h); protection by ent-5β,3α-A also persisted longer (~3 h) than for 5β,3α-A (~2 h).

Conclusions

The unnatural enantiomers of 17-keto androgen class neurosteroids have greater in vivo potency and a longer duration of action than their natural counterparts. The more prolonged duration of action of the unnatural enantiomers could reflect reduced susceptibility to metabolism. Unnatural enantiomers of androgen class neurosteroids could have therapeutic utility and may provide advantages over the corresponding natural isomers due to enhanced potency and improved pharmacokinetic characteristics.     

Characterization of neurosteroid effects on hyperpolarizing current at α4β2δ GABAA receptors

Rationale

The neurosteroid 3α,5β-THP (3α-OH-5β-pregnan-20-one, pregnanolone) is a modulator of the GABA_A receptor (GABAR), with α4β2δ GABARs the most sensitive. However, the effects of 3α,5β-THP at α4β2δ are polarity-dependent: 3α,5β-THP potentiates depolarizing current, as has been widely reported, but decreases hyperpolarizing current by accelerating desensitization.

Objectives

The present study further characterized 3α,5β-THP inhibition of hyperpolarizing current at this receptor and compared effects of other related steroids at α4β2δ GABARs.

Methods

α4β2δ GABARs were expressed in HEK-293 cells, and agonist-gated current recorded with whole cell voltage-clamp techniques using a theta tube to rapidly apply agonist before and after application of neurosteroids.

Results

The GABA-modulatory steroids (30 nM) 3α,5α-THP (3α-OH-5α-pregnan-20-one, allopregnanolone) and THDOC (3α,21-dihydroxy-5α-pregnan-20-one) inhibited hyperpolarizing GABA (10 μM)-gated current at α4β2δ GABARs similar to 3α,5β-THP, while the inactive 3β,5β-THP isomer had no effect. Greater inhibition was seen for current gated by the high efficacy agonist gaboxadol (THIP, 100 μM) than for GABA (0.1–1000 μM), consistent with an effect of 3α,5β-THP on desensitization. Inhibitory effects of the steroid were not seen under low [Cl^?] conditions or in the presence of calphostin C (500 nM), an inhibitor of protein kinase C. Chimeras swapping the IL (intracellular loop) of α4 with α1, when expressed with β2 and δ, produced receptors (α[414]β2δ) which were not inhibited by 3α,5β-THP when GABA-gated current was hyperpolarizing, while α[141]β2δ exhibited steroid-induced polarity-dependent modulation.

Conclusions

These findings suggest that numerous neurosteroids exhibit polarity-dependent effects at α4β2δ GABARs, which are dependent upon protein kinase C and the IL of α4.     

Involvement of insular muscarinic cholinergic receptors in morphine-induced conditioned place preference in rats

Rationale

Drug addiction represents a pathological usurpation of neural processes involved in learning and memory. Retrieval of drug-related memories can result in drug craving and relapse. Recently, the insula was identified as part of the neuronal circuit responsible for the processing of drug memory; however, its precise role remains unclear.

Objective

To investigate the involvement of insular muscarinic acetylcholine receptors (mAChRs) in the processing of drug memory.

Method

The morphine-induced conditioned place preference (CPP) was used to assess drug memory. All rats were first trained with morphine to establish the CPP. Sub-groups of these rats were used for contextual cue-induced CPP reinstatement. Other sub-groups of rats underwent extinction of the CPP, and 5 m/kg morphine was used for priming-induced CPP reinstatement. Microinjection of mAChR antagonists or agonists into the insula was performed prior to the CPP tests in order to evaluate their effect on CPP expression.

Results

Insular microinjections of the nonselective mAChR antagonist, scopolamine, and the M₁ antagonist, pirenzepine, significantly inhibited CPP expression in both contextual cue- and priming-induced CPP reinstatement; the M₁ agonist, MCN-A-343, and the M₄ antagonist, tropicamide, enhanced CPP expression. The M₄ agonist, LY2033298, inhibited CPP expression. The M₂ antagonist, methoctramine, and M₃ antagonist, 4-DAMP, had no effect on CPP expression.

Conclusion

Our results demonstrate that insular mAChRs play a role in the processing of drug memory. M₁ and M₄ mAChRs work paradoxically; M₁ activation and M₄ inhibition attenuate the expression of drug memory, while M₁ inhibition and M₄ activation augment the expression of drug memory.     

Neuroprotection by the synthetic neurosteroid enantiomers ent-PREGS and ent-DHEAS against Aβ25–35 peptide-induced toxicity in vitro and in vivo in mice

Rationale

Pregnenolone sulfate (PREGS) and dehydroepiandrosterone sulphate (DHEAS) are pro-amnesic, anti-amnesic and neuroprotective steroids in rodents. In Alzheimer’s disease (AD) patient’s brains, their low concentrations are correlated with high levels of Aβ and tau proteins. The unnatural enantiomer ent-PREGS enhanced memory in rodents. We investigated here whether ent-PREGS and ent-DHEAS could be neuroprotective in AD models.

Objective

The effects of PREGS, ent-PREGS, DHEAS and ent-DHEAS against Aβ_25–35 peptide-induced toxicity were examined in vitro on B104 neuroblastoma cells and in vivo in mice.

Methods

B104 cells pretreated with the steroids before Aβ_25–35 were analysed by flow cytometry measuring cell viability and death processes. Mice injected intracerebroventricularly with Aβ_25–35 and the steroids were analysed for their memory abilities. Additionally, lipid peroxidation levels in the hippocampus were measured.

Results

ent-PREGS and PREGS significantly attenuated the Aβ_25–35-induced decrease in cell viability. Both steroids prevented the Aβ_25–35-induced increase in late apoptotic cells. PREGS further attenuated the ratio of necrotic cells. ent-DHEAS and DHEAS significantly reduced the Aβ_25–35-induced toxicity and prevented the cells from entering late apoptosis and necrosis. All steroids stimulated neurite outgrowth per se and prevented the Aβ_25–35-induced decrease. In vivo, ent-PREGS and ent-DHEAS significantly attenuated the Aβ_25–35-induced decrease in memory (spontaneous alternation and passive avoidance) and an increase in lipid peroxidation levels. In contrast to the natural steroids, both enantiomers prevented amnesia when injected 6 h before Aβ_25–35 in contrast to the natural steroids.

Conclusion

The unnatural steroids ent-PREGS and ent-DHEAS are potent neuroprotective agents and could be effective therapeutical tools in AD.     

Low anandamide doses facilitate male rat sexual behaviour through the activation of CB1 receptors

Rationale

Endocannabinoids (eCN) exert biphasic effects on several behaviours; however, they have only been reported to inhibit male sexual behaviour. eCN, the endogenous ligands for CB1 receptors, are released in response to neuronal stimulation and regulate the functioning of the mesocorticolimbic system (MCL), which is activated by male sexual behaviour. We hypothesised that eCN might exert biphasic effects on male rat copulatory behaviour and be released during copulation to satiation as a result of the repeated activation of the MCL system.

Objectives

The study was conducted to determine if the eCN anandamide exerts biphasic effects on sexual behaviour expression of sexually experienced and sexually satiated male rats and to establish the possible participation of eCN in the sexual satiation phenomenon.

Methods

The eCN anandamide and the CB₁ receptor antagonist AM251 were systemically administered to sexually experienced or sexually satiated rats and their effects on copulatory behaviour analysed.

Results

Low anandamide doses facilitated sexual behaviour expression in sexually experienced and in sexually satiated rats by acting at CB₁ receptors. AM251 blocked the establishment of the sexual inhibition that characterises sexual satiation, but did not reverse it once established.

Conclusions

Anandamide exerts dose-based biphasic effects on copulatory behaviour of sexually experienced male rats and facilitates sexual behaviour expression of sexually satiated animals at low doses. eCN participate in the establishment, but not in the maintenance of the sexual inhibitory state that characterises the sexual satiation phenomenon.     

Opposing neural effects of naltrexone on food reward and aversion: implications for the treatment of obesity

Rationale

Opioid antagonism reduces the consumption of palatable foods in humans but the neural substrates implicated in these effects are less well understood.

Objectives

The aim of the present study was to examine the effects of the opioid antagonist, naltrexone, on neural response to rewarding and aversive sight and taste stimuli.

Methods

We used functional magnetic resonance imaging (fMRI) to examine the neural responses to the sight and taste of pleasant (chocolate) and aversive (mouldy strawberry) stimuli in 20 healthy volunteers who received a single oral dose of naltrexone (50 mg) and placebo in a double-blind, repeated-measures cross-over, design.

Results

Relative to placebo, naltrexone decreased reward activation to chocolate in the dorsal anterior cingulate cortex and caudate, and increased aversive-related activation to unpleasant strawberry in the amygdala and anterior insula.

Conclusions

These findings suggest that modulation of key brain areas involved in reward processing, cognitive control and habit formation such as the dorsal anterior cingulate cortex (dACC) and caudate might underlie reduction in food intake with opioid antagonism. Furthermore we show for the first time that naltrexone can increase activations related to aversive food stimuli. These results support further investigation of opioid treatments in obesity.     

A Floating Hydrogel System Capable of Generating CO2 Bubbles to Diminish Urinary Obstruction After Intravesical Instillation

Aim

Intravesical instillation is commonly used to decrease the tumor recurrence after transurethral resection. However, most drug solutions would be eliminated from bladder after the first voiding of urine, so its clinical efficacy is limited. To overcome this obstacle, we developed a floating hydrogel system for controlled delivery of antitumor drugs.

Methods

The floating hydrogel was made of Adriamycin, thermo-sensitive polymer (Poloxamer 407) and NH₄HCO₃, which was liquid at low temperature while forming hydrogel at high temperature. Meanwhile, at high temperature, NH₄HCO₃ decomposed to produce CO₂ bubbles, which helped hydrogel float in bladder without urinary obstruction.

Results

The mixture containing 45% P407 and 6% NH₄HCO₃ was selected as the optimal formulation. At 37°C, the mixture formed hydrogel and produced many bubbles which could be observed by B ultrasound. The vitro study showed that the antitumor drug Doxorubicin was released in a controlled manner. After the mixture was instilled into rabbit bladder, it formed hydrogel and floated in the bladder. The bladder stimuli was reduced and antitumor drugs could be released continuously in the bladder.

Conclusion

Our results suggested that the floating hydrogel was a feasible intravesical drug delivery system and may have application prospects in intravesical therapy for bladder cancer.     

Beta-2 adrenergic receptors mediate stress-evoked reinstatement of cocaine-induced conditioned place preference and increases in CRF mRNA in the bed nucleus of the stria terminalis in mice

Rationale

Understanding the mechanisms responsible for stress-induced relapse is important for guiding treatment strategies aimed at minimizing the contribution of stress to addiction. Evidence suggests that these mechanisms involve interactions between noradrenergic systems and the neuropeptide corticotropin-releasing factor (CRF).

Objectives

The interaction between β-adrenergic receptors (ARs) and CRF as it relates to the reinstatement of cocaine-conditioned reward in response to a stressor was examined in mice. We hypothesized that β₂-ARs are required for stress-induced activation of CRF pathways responsible for reinstatement.

Methods

Stress-induced relapse was examined based on the re-establishment of cocaine-induced conditioned place preference (CPP; 4?×?15 mg/kg cocaine, i.p.) after extinction using forced swim (6 min at 22 °C) or an injection of the β₂-AR agonist, clenbuterol (4 mg/kg, i.p.). The CRF-R1 antagonist antalarmin (10 mg/kg, i.p.) or the β₂-AR antagonist ICI-118,551 (1 mg/kg, i.p.) were given 30 min prior to reinstating stimuli. Quantitative PCR was conducted in dissected bed nucleus of the stria terminalis (BNST) and amygdala, putative sources of CRF that contribute to reinstatement, to examine the effects of ICI-118,551 on swim-induced increases in CRF messenger RNA (mRNA) in mice with a cocaine history.

Results

Pretreatment with ICI-118,551 or antalarmin blocked swim-induced reinstatement of CPP. Reinstatement by clenbuterol was also blocked by antalarmin. ICI-118,551 pretreatment prevented swim-induced increases in CRF mRNA in the BNST. Effects in the amygdala were not observed.

Conclusions

These findings indicate that, during stress, norepinephrine, via β₂-ARs, either directly or indirectly activates CRF-releasing neurons in the BNST that interface with motivational neurocircuitry to induce reinstatement of cocaine-conditioned reward.     

New Thermoresistant Polymorph from CO2 Recrystallization of Minocycline Hydrochloride

Purpose

To prepare and thoroughly characterize a new polymorph of the broad-spectrum antibiotic minocycline from its hydrochloride dehydrate salts.

Methods

The new minocycline hydrochloride polymorph was prepared by means of the antisolvent effect caused by carbon dioxide. Minocycline recrystallized as a red crystalline hydrochloride salt, starting from solutions or suspensions containing CO₂ and ethanol under defined conditions of temperature, pressure and composition.

Results

This novel polymorph (β-minocycline) revealed characteristic PXRD and FTIR patterns and a high melting point (of 247?ºC) compared to the initial minocycline hydrochloride hydrates (α-minocycline). Upon dissolution the new polymorph showed full anti-microbial activity. Solid-state NMR and DSC studies evidenced the higher chemical stability and crystalline homogeneity of β-minocycline compared to the commercial chlorohydrate powders. Molecular structures of both minocyclines present relevant differences as shown by multinuclear solid-state NMR.

Conclusions

This work describes a new crystalline structure of minocycline and evidences the ability of ethanol-CO₂ system in removing water molecules from the crystalline structure of this API, at modest pressure, temperature and relatively short time (2 h), while controlling the crystal habit. This process has therefore the potential to become a consistent alternative towards the control of the solid form of APIs.     

Effects of oxycodone on brain responses to emotional images

Rationale

Evidence from animal and human studies suggests that opiate drugs decrease emotional responses to negative stimuli and increase responses to positive stimuli. Such emotional effects may motivate misuse of oxycodone (OXY), a widely abused opiate. Yet, we know little about how OXY affects neural circuits underlying emotional processing in humans.

Objective

We examined effects of OXY on brain activity during presentation of positive and negative visual emotional stimuli. We predicted that OXY would decrease amygdala activity to negative stimuli and increase ventral striatum (VS) activity to positive stimuli. Secondarily, we examined the effects of OXY on other emotional network regions on an exploratory basis.

Methods

In a three-session study, healthy adults (N?=?17) received placebo, 10 and 20 mg OXY under counterbalanced, double-blind conditions. At each session, participants completed subjective and cardiovascular measures and underwent functional MRI (fMRI) scanning while completing two emotional response tasks.

Results

Our emotional tasks reliably activated emotional network areas. OXY produced subjective effects but did not alter either behavioral responses to emotional stimuli or activity in our primary areas of interest. OXY did decrease right medial orbitofrontal cortex (MOFC) responses to happy faces.

Conclusions

Contrary to our expectations, OXY did not affect behavioral or neural responses to emotional stimuli in our primary areas of interest. Further, the effects of OXY in the MOFC would be more consistent with a decrease in value for happy faces. This may indicate that healthy adults do not receive emotional benefits from opiates, or the pharmacological actions of OXY differ from other opiates.     

Effect of varenicline on aspects of inhibitory control in smokers

Rationale

Varenicline, a partial agonist at α4β2 nicotinic receptors (nAChRs) aids smoking cessation by reducing craving. Successful quitting may be associated with greater inhibitory control but the effectiveness of varenicline in this regard is unknown.

Objectives

This study aimed to investigate the effect of varenicline on aspects of inhibitory control in smokers.

Methods

A double-blind, placebo-controlled study investigating the effect of varenicline 1 mg (or matched placebo) in satiated and abstinent smokers. Tests included Rapid Visual Information Processing (RVIP), Stop-Signal (SS), Prospective Memory (PM) and the Cambridge Gambling Task (CGT).

Results

Smoking enhanced RVIP accuracy and latency to respond. Varenicline did not alter RVIP performance, nor the effect of smoking, suggesting that these effects were unrelated to α4β2 nAChRs. Smoking increased the number of errors during SS and increased the stop latency, indicating that smoking decreased inhibitory control. Varenicline partially mimicked this effect of smoking but also reduced the smoking-induced increase, indicating a role for α4β2 nAChRs. Likewise, smoking increased the number of points bet following a win during CGT and varenicline blocked this effect. There was no effect of smoking or varenicline on PM target detection per se. However, smoking protected the target detection rate in the ongoing task when a concurrent intention was introduced. Varenicline improved response speed in both satiated and abstinent smokers.

Conclusions

Some aspects of inhibitory control may be mediated by α4β2-related mechanisms and blockade of smoking-induced disinhibition may contribute towards the action of varenicline as an aid to smoking cessation.     

Disruption of latent inhibition induced by ovariectomy can be reversed by estradiol and clozapine as well as by co-administration of haloperidol with estradiol but not by haloperidol alone

Introduction

Epidemiological and clinical life cycle studies have indicated that the more favorable illness course and the better response to antipsychotic drugs (APDs) in women with schizophrenia correlate with high levels of estrogen, whereas increased vulnerability to exacerbation and relapse and reduced sensitivity to treatment are associated with low estrogen levels. Accordingly, the estrogen hypothesis of schizophrenia proposes that estrogen has a neuroprotective effect in women vulnerable to schizophrenia.

Materials and methods

Latent inhibition (LI), the capacity to ignore stimuli that received nonreinforced preexposure prior to conditioning, is disrupted in acute schizophrenia patients and in rats and humans treated with the psychosis inducing drug amphetamine. Disruption of LI is reversible by typical and atypical APDs. The present study tested whether low levels of estrogen induced by ovariectomy (OVX) would lead to disruption of LI in female rats and whether such disruption would be normalized by estrogen replacement treatment and/or APDs.

Results

Results showed that OVX led to LI disruption, which was reversed by 17β-estradiol (150 μg/kg) and the atypical APD clozapine (5 mg/kg), but not by the typical APD haloperidol (0.1, 0.2, 0.3 mg/kg). Haloperidol regained efficacy when administered with 17β-estradiol (50 μg/kg).

Discussion

These results provide the first demonstration in rats that low levels of hormones can induce a pro-psychotic state that is resistant to at least typical antipsychotic treatment. This constellation may mimic states seen in schizophrenic women during periods associated with low levels of hormones such as the menopause.     

Fast-Acting Clotrimazole Composited PVP/HPβCD Nanofibers for Oral Candidiasis Application

Purpose

This study investigates fabrication of clotrimazole (CZ)-composited electrospun Polyvinylpyrrolidone/Hydroxypropyl-β-cyclodextrin (PVP/HPβCD) blended nanofiber mats for oral candidiasis applications.

Methods

PVP/HPβCD blended nanofiber mats containing clotrimazole were electrospun and characterized using SEM, DSC and XRPD. The solvent system ethanol: water: benzyl alcohol (EtOH:H₂O:BzOH) with a 70:20:10 ratio was optimal for the electrospinning process. Various amounts of CZ were loaded into the nanofiber mats. The nanofiber mats was further investigated for drug release, antifungal activity and cytotoxicity.

Results

The fiber diameters in the mats were in the nanometer range. The DSC and XRPD revealed a molecular dispersion of amorphous CZ in the nanofiber mats. The loading capacity increased when CZ content was raised. A fast dissolved and released of CZ from the nanofibers mat was achieved. The ability of the CZ-loaded nanofiber mats to kill the Candida depended on the amount of CZ in the mats; moreover, the CZ-loaded nanofibers killed the Candida significantly faster than the CZ powder and lozenges with low cytotoxicity.

Conclusions

CZ-loaded nanofiber mats were successfully electrospun. They exhibited rapid antifungal activity in vitro relative to CZ powder and lozenges. Further in vivo studies are needed to investigate for their application in oral candidiasis.     

Effects of coincident 5-HT1A receptor stimulation and NMDA receptor antagonism on l-DOPA-induced dyskinesia and rotational behaviors in the hemi-parkinsonian rat

Rationale

Serotonin 1A receptor (5-HT1AR) agonists reduce l-DOPA-induced dyskinesia and enhance motor function in experimental and clinical investigations of Parkinson’s disease (PD). While the mechanism(s) by which these effects occur are unclear, recent research suggests that modulation of glutamate neurotransmission contributes.

Objective

To further delineate the relationship between 5-HT_1A receptors and glutamate, the current study examined the effects of the 5-HT_1AR agonist, ±8-OH-DPAT and the N-methyl-d-aspartic acid receptor (NMDAR) antagonist, MK-801, on l-DOPA-induced motor behavior.

Materials and methods

Unilateral 6-hydroxydopamine lesioned male Sprague–Dawley rats were rendered dyskinetic with 1 week of daily l-DOPA (12 mg/kg, i.p.) + benserazide (15 mg/kg, i.p.). On test days, one group of rats received pretreatments of: ±8-OH-DPAT (0, 0.03, 0.1, 0.3 mg/kg, i.p.) or MK-801 (0, 0.03, 0.1, 0.3 mg/kg, i.p.). A second group was administered combined ±8-OH-DPAT (0, 0.03 or 0.1 mg/kg, i.p.) + MK-801 (0, 0.1 mg/kg, i.p.). Pretreatments were followed by l-DOPA administration, after which, abnormal involuntary movements (AIMs) and rotations were monitored. To investigate effects on motor performance, subthreshold doses of ±8-OH-DPAT (0.03 mg/kg, i.p.) + MK-801 (0.1 mg/kg, i.p.) were administered to l-DOPA-naïve hemiparkinsonian rats before the forepaw adjusting steps test.

Results

Individually, both ±8-OH-DPAT and MK-801 dose-dependently decreased l-DOPA-induced AIMs without affecting rotations. Combined subthreshold doses of ±8-OH-DPAT+MK-801 reduced l-DOPA-induced AIMs and potently enhanced contralateral rotations without altering l-DOPA-induced motor improvements.

Conclusions

The current results indicate a functional interaction between 5-HT_1AR and NMDAR that may improve pharmacological treatment of PD patients.