Circulating Pro-Inflammatory Exosomes Worsen Stroke Outcomes in Aging.

[Figure: see text].

Systemic Administration of α7-Nicotinic Acetylcholine Receptor Ligands Does Not Improve Renal Injury or Behavior in Mice With Advanced Systemic Lupus Erythematosus.

There is a critical need for safe treatment options to control inflammation in patients with systemic lupus erythematosus (SLE) since the inflammation contributes to morbidity and mortality in advanced disease. Endogenous neuroimmune mechanisms like the cholinergic anti-inflammatory pathway can be targeted to modulate inflammation, but the ability to manipulate such pathways and reduce inflammation and end organ damage has not been fully explored in SLE. Positive allosteric modulators (PAM) are pharmacological agents that inhibit desensitization of the nicotinic acetylcholine receptor (α7-nAChR), the main anti-inflammatory feature within the cholinergic anti-inflammatory pathway, and may augment α7-dependent cholinergic tone to generate therapeutic benefits in SLE. In the current study, we hypothesize that activating the cholinergic anti-inflammatory pathway at the level of the α7-nAChR with systemic administration of a partial agonist, GTS-21, and a PAM, PNU-120596, would reduce inflammation, eliminating the associated end organ damage in a mouse model of SLE with advanced disease. Further, we hypothesize that systemic α7 ligands will have central effects and improve behavioral deficits in SLE mice. Female control (NZW) and SLE mice (NZBWF1) were administered GTS-21 or PNU-120596 subcutaneously via minipumps for 2 weeks. We found that the increased plasma dsDNA autoantibodies, splenic and renal inflammation, renal injury and hypertension usually observed in SLE mice with advanced disease at 35 weeks of age were not altered by GTS-21 or PNU-120596. The anxiety-like behavior presented in SLE mice was also not improved by GTS-21 or PNU-120596. Although no significant beneficial effects of α7 ligands were observed in SLE mice at this advanced stage, we predict that targeting this receptor earlier in the pathogenesis of the disease may prove to be efficacious and should be addressed in future studies.

Therapeutic efficacy of α7 ligands after acute ischaemic stroke is linked to conductive states of α7 nicotinic ACh receptors.

BACKGROUND AND PURPOSE: Targeting α7 nicotinic ACh receptors (nAChRs) in neuroinflammatory disorders including acute ischaemic stroke holds significant therapeutic promise. However, therapeutically relevant signalling mechanisms remain unidentified. Activation of neuronal α7 nAChRs triggers ionotropic signalling, but there is limited evidence for it in immunoglial tissues. The α7 ligands which are effective in reducing acute ischaemic stroke damage promote α7 ionotropic activity, suggesting a link between their therapeutic effects for treating acute ischaemic stroke and activation of α7 conductive states. EXPERIMENTAL APPROACH: This hypothesis was tested using a transient middle cerebral artery occlusion (MCAO) model of acute ischaemic stroke, NS6740, a known selective non-ionotropic agonist of α7 nAChRs and 4OH-GTS-21, a partial α7 agonist. NS6740-like ligands exhibiting low efficacy/potency for ionotropic activity will be referred to as non-ionotropic agonists or "metagonists". KEY RESULTS: 4OH-GTS-21, used as a positive control, significantly reduced neurological deficits and brain injury after MCAO as compared to vehicle and NS6740. By contrast, NS6740 was ineffective in identical assays and reversed the effects of 4OH-GTS-21 when these compounds were co-applied. Electrophysiological recordings from acute hippocampal slices obtained from NS6740-injected animals demonstrated its remarkable brain availability and protracted effects on α7 nAChRs as evidenced by sustained (>8 h) alterations in α7 ionotropic responsiveness. CONCLUSION AND IMPLICATIONS: These results suggest that α7 ionotropic activity may be obligatory for therapeutic efficacy of α7 ligands after acute ischaemic stroke yet, highlight the potential for selective application of α7 ligands to disease states based on their mode of receptor activation.

Spleen is not required for therapeutic effects of 4OH-GTS-21, a selective α7 nAChR agonist, in the sub-acute phase of ischemic stroke in rats.

Acute ischemic stroke (AIS) causes both central and peripheral inflammation, while activation of α7 nicotinic acetylcholine receptors (nAChRs) provides both central and peripheral anti-inflammatory and anti-apoptotic effects. Here, we provide evidence that 4OH-GTS-21, a selective α7 agonist, produces its therapeutic effects via primarily central sites of action because 4OH-GTS-21 was found equally effective in splenectomized and non-spenectomized rats in the sub-acute phase of ischemic stroke (≤1 week). However, the spleen may boost the therapeutic efficacy of 4OH-GTS-21 in certain behavioral tasks as our data also indicated. In our tests, AIS was modeled by transient middle cerebral artery occlusion (tMCAO). Splenectomy was done 2 weeks before tMCAO. We determined that: 1) Daily 4OH-GTS-21 treatments for 7 days after tMCAO significantly reduced neurological deficits and brain injury in both splenectomized and non-spelenectomized rats demonstrating that the spleen is not required for therapeutic benefits of 4OH-GTS-21; 2) The effects of 4OH-GTS-21 in the adhesive sticker removal test were significantly weaker in splenectomized animals suggesting that the spleen boosts the efficacy of 4OH-GTS-21 in the first week after tMCAO; and 3) Ischemic brain injury was not significantly affected by splenectomy in both vehicle-treated and 4OH-GTS-21-treated animals. These data support the hypothesis that the therapeutic efficacy of sub-chronic (≤1 week) 4OH-GTS-21 primarily originates from central sites of action. These results validate brain availability as a critical factor for developing novel α7 ligands for AIS.

Expression and Function of Transient Receptor Potential Ankyrin 1 Ion Channels in the Caudal Nucleus of the Solitary Tract.

The nucleus of the solitary tract (NTS) receives visceral information via the solitary tract (ST) that comprises the sensory components of the cranial nerves VII, IX and X. The Transient Receptor Potential Ankyrin 1 (TRPA1) ion channels are non-selective cation channels that are expressed primarily in pain-related sensory neurons and nerve fibers. Thus, TRPA1 expressed in the primary sensory afferents may modulate the function of second order NTS neurons. This hypothesis was tested and confirmed in the present study using acute brainstem slices and caudal NTS neurons by RT-PCR, immunostaining and patch-clamp electrophysiology. The expression of TRPA1 was detected in presynaptic locations, but not the somata of caudal NTS neurons that did not express TRPA1 mRNA or proteins. Moreover, caudal NTS neurons did not show somatodendritic responsiveness to TRPA1 agonists, while TRPA1 immunostaining was detected only in the afferent fibers. Electrophysiological recordings detected activation of presynaptic TRPA1 in glutamatergic terminals synapsing on caudal NTS neurons evidenced by the enhanced glutamatergic synaptic neurotransmission in the presence of TRPA1 agonists. The requirement of TRPA1 for modulation of spontaneous synaptic activity was confirmed using TRPA1 knockout mice where TRPA1 agonists failed to alter synaptic efficacy. Thus, this study provides the first evidence of the TRPA1-dependent modulation of the primary afferent inputs to the caudal NTS. These results suggest that the second order caudal NTS neurons act as a TRPA1-dependent interface for visceral noxious-innocuous integration at the level of the caudal brainstem.

Treatment duration affects cytoprotective efficacy of positive allosteric modulation of α7 nAChRs after focal ischemia in rats.

To minimize irreversible brain injury after acute ischemic stroke (AIS), the time to treatment (i.e., treatment delay) should be minimized. However, thus far, all cytoprotective clinical trials have failed. Analysis of literature identified short treatment durations (≤72 h) as a common motif among completed cytoprotective clinical trials. Here, we argue that short cytoprotective regimens even if given early after AIS may only slow down the evolution of ischemic brain injury and fail to deliver sustained long-term solutions leading to relapses that may be misinterpreted for conceptual failure of cytoprotection. In this randomized blinded study, we used young adult male rats subjected to transient 90 min suture middle cerebral artery occlusion (MCAO) and treated with acute vs. sub-chronic regimens of PNU120596, a prototypical positive allosteric modulator of α7 nicotinic acetylcholine receptors with anti-inflammatory cytoprotective properties to test the hypothesis that insufficient treatment durations may reduce therapeutic benefits of otherwise efficacious cytoprotectants after AIS. A single acute treatment 90 min after MCAO significantly reduced brain injury and neurological deficits 24 h later, but these effects vanished 72 h after MCAO. These relapses were avoided by utilizing sub-chronic treatments. Thus, extending treatment duration augments therapeutic efficacy of PNU120596 after MCAO. Furthermore, sub-chronic treatments could offset the negative effects of prolonged treatment delays in cases where the acute treatment window after MCAO was left unexploited. We conclude that a combination of short treatment delays and prolonged treatment durations may be required to maximize therapeutic effects of PNU120596, reduce relapses and ensure sustained therapeutic efficacy after AIS. Similar concepts may hold for other cytoprotectants including those that failed in clinical trials.

Duration of isoflurane-based surgical anesthesia determines severity of brain injury and neurological deficits after a transient focal ischemia in young adult rats.

Tremendous efforts and funds invested in discovery of novel drug treatments for ischemic stroke have so far failed to deliver clinically efficacious therapies. The reasons for these failures are not fully understood. An indiscriminate use of isoflurane-based surgical anesthesia with or without nitrous oxide may act as an unconstrained, untraceable source of data variability, potentially causing false-positive or false-negative results. To test this hypothesis, a common transient suture middle cerebral artery occlusion (tMCAO) model of ischemic stroke in young adult male rats was used to determine the impact of a typical range of anesthesia durations required for this model on data variability (i.e., infarct volume and neurological deficits). The animals were maintained on spontaneous ventilation. The study results indicated that: (1) Variable duration of isoflurane anesthesia prior, during and after tMCAO is a significant source of data variability as evidenced by measurements of infarct volume and neurological deficits; and (2) Severity of brain injury and neurological deficits after tMCAO is inversely related to the duration of isoflurane anesthesia: e.g., in our study, a 90min isoflurane anesthesia nearly completely protected brain tissues from tMCAO-induced injury and thus, would be expected to obscure the effects of stroke treatments in pre-clinical trials. To elevate transparency, rigor and reproducibility of stroke research and minimize undesirable effects of isoflurane on the outcome of novel drug testing, we propose to monitor, minimize and standardize isoflurane anesthesia in experimental surgeries and make anesthesia duration a required reportable parameter in pre-clinical studies. Specifically, we propose to adopt 20-30min as an optimal anesthesia duration that both minimizes neuroprotective effects of isoflurane and permits a successful completion of surgical procedures in a suture tMCAO model of ischemic stroke in rodents. As the mechanisms and neuroprotective, metabolic and immune effects of general anesthesia are not fully understood, the results of this study cannot be blindly generalized to other anesthetics, animal species and experimental models.

Boosting Endogenous Resistance of Brain to Ischemia.

Most survivors of ischemic stroke remain physically disabled and require prolonged rehabilitation. However, some stroke victims achieve a full neurological recovery suggesting that the human brain can defend itself against ischemic injury, but the protective mechanisms are unknown. This study used selective pharmacological agents and a rat model of cerebral ischemic stroke to detect endogenous brain protective mechanisms that require activation of α7 nicotinic acetylcholine receptors (nAChRs). This endogenous protection was found to be (1) limited to less severe injuries; (2) significantly augmented by intranasal administration of a positive allosteric modulator of α7 nAChRs, significantly reducing brain injury and neurological deficits after more severe ischemic injuries; and (3) reduced by inhibition of calcium/calmodulin-dependent kinase-II. The physiological role of α7 nAChRs remains largely unknown. The therapeutic activation of α7 nAChRs after cerebral ischemia may serve as an important physiological responsibility of these ubiquitous receptors and holds a significant translational potential.

Allosteric Modulation of Nicotinic Acetylcholine Receptors: The Concept and Therapeutic Trends.

Expressing functional nicotinic acetylcholine receptors (nAChRs) may be beneficial to central neurons and neuronal networks because activation of nAChRs enhances neuronal resistance to injury, improves attention, cognitive performance, and produces robust anti-inflammatory and analgesic effects in mammals. Although exogenous orthosteric nAChR ligands present valuable tools in treatment of age- and trauma-related neurological deficits, therapeutic approaches that could amplify the brain's innate ability to maintain cholinergic homeostasis and resist injury may serve as intriguing and promising alternatives and have not been fully explored. One of these novel approaches utilizes positive allosteric modulators (PAMs) of nAChRs. Because of the ubiquitous expression of nAChRs in neuronal, glial and immune tissues, highly selective PAMs could amplify multiple endogenous neuroprotective, pro-cognitive, anti-inflammatory and anti-nociceptive cholinergic pathways to offset cholinergic hypofunction and generate therapeutic efficacy by targeting only a single player: i.e., nAChRs activated by endogenous cholinergic tone. In this article, I review the concept of allosteric modulation and current trends in therapeutic applications of nicotinic PAMs.

Targeting the Cholinergic System for Neuroprotection and/or Enhancement of Functional Recovery Following Neurotrauma.

Development of novel pharmacotherapies for the treatment of traumatic injury to the nervous system has been ongoing for over 40 years. Despite many promising compounds discovered using animal models, no treatments have successfully translated into the clinic. The central dogma in this field is that brain trauma initiates a complex chain of biochemical events leading to secondary brain damage and sustained neurological deficits. The delayed secondary brain injury is likely to result from multiple insults including oxidative stress, mitochondrial dysfunction, breakdown of the blood brain barrier, dysregulated release of glutamate, pro-inflammatory cytokines, and other mediators. However, therapies targeting these systems have generally met with failure in clinical trials. The purpose of this review is to summarize the models used for preclinical neurotrauma research, provide a brief overview of previous failed clinical trials in head and spinal cord injury, and finally, to review involvement of the cholinergic system and discuss implications for future research. Possibilities and pitfalls of targeting the cholinergic system for neuroprotection and/or enhancement of functional recovery are also discussed.

The Cholinergic Potential, the Vagus Nerve and Challenges in Treatment of Traumatic Brain Injury.

Existing treatments of traumatic brain injury (TBI) have failed to reverse tremendous losses in productivity, independence and overall quality of life among TBI victims and therefore, cannot be viewed as sufficient. Although there is no shortage of promising basic concepts that may translate to efficacious therapies after TBI, the accumulated knowledge has yet to deliver treatments that adequately meet clinical and social demands. In this article, we discuss novel concepts, recent advances and accompanying challenges in developing cholinergic and related therapies after TBI.

High therapeutic potential of positive allosteric modulation of α7 nAChRs in a rat model of traumatic brain injury: proof-of-concept.

There are currently no clinically efficacious drug therapies to treat brain damage secondary to traumatic brain injury (TBI). In this proof-of-concept study, we used a controlled cortical impact model of TBI in young adult rats to explore a novel promising approach that utilizes PNU-120596, a previously reported highly selective Type-II positive allosteric modulator (α7-PAM) of α7 nicotinic acetylcholine receptors (nAChRs). α7-PAMs enhance and prolong α7 nAChR activation, but do not activate α7 nAChRs when administered without an agonist. The rational basis for the use of an α7-PAM as a post-TBI treatment is tripartite and arises from: (1) the intrinsic ability of brain injury to elevate extracellular levels of choline (a ubiquitous cell membrane-building material and a selective endogenous agonist of α7 nAChRs) due to the breakdown of cell membranes near the site and time of injury; (2) the ubiquitous expression of functional α7 nAChRs in neuronal and glial/immune brain cells; and (3) the potent neuroprotective and anti-inflammatory effects of α7 nAChR activation. Therefore, both neuroprotective and anti-inflammatory effects can be achieved post-TBI by targeting only a single player (i.e., the α7 nAChR) using α7-PAMs to enhance the activation of α7 nAChRs by injury-elevated extracellular choline. Our data support this hypothesis and demonstrate that subcutaneous administration of PNU-120596 post-TBI in young adult rats significantly reduces both brain cell damage and reactive gliosis. Therefore, our results introduce post-TBI systemic administration of α7-PAMs as a promising therapeutic intervention that could significantly restrict brain injury post-TBI and facilitate recovery of TBI patients.

Nicotine enhances inhibition of mouse vagal motor neurons by modulating excitability of premotor GABAergic neurons in the nucleus tractus solitarii.

The caudal nucleus of the solitary tract (NTS) serves as the site of the first synapse for visceral sensory inputs to the central nervous system. The NTS sends functional projections to multiple brain nuclei, with gastric-related projections primarily targeting the dorsal motor nucleus of the vagus (DMV). Previous studies have demonstrated that the majority of caudal NTS neurons that project to the DMV respond robustly to nicotine and express nicotinic acetylcholine receptors (nAChRs). However, the cytochemical identity and relationship with specific viscera of DMV-projecting, nicotine-responsive caudal NTS neurons have not been determined. The present study used transgenic mice that express enhanced green fluorescent protein (EGFP) under a GAD67 promoter in a subset of GABAergic neurons, in vivo retrograde pseudorabies viral labeling to identify gastric-related vagal complex neurons, and patch-clamp electrophysiology in acute brain stem slices to test the hypothesis that gastric-related and GABAergic inhibitory synaptic input to the DMV from the caudal NTS is under a robust modulatory control by nAChRs. Our results suggest that activation of nAChRs in the caudal NTS, but not DMV, potentiates GABAergic, but not glutamatergic, input to the DMV. Gastric-related caudal NTS and DMV neurons are directly involved in this nicotine-sensitive circuitry. Understanding the central patterns of nicotinic modulation of visceral sensory-motor circuitry may help develop therapeutic interventions to restore autonomic homeostasis in patients with autonomic impairments.

Cerebrospinal Fluid Levels of Monoamine Metabolites in the Epileptic Baboon.

The baboon represents a natural model for genetic generalized epilepsy and sudden unexpected death in epilepsy (SUDEP). In this retrospective study, cerebrospinal fluid (CSF) monoamine metabolites and scalp electroencephalography (EEG) were evaluated in 263 baboons of a pedigreed colony. CSF monoamine abnormalities have been linked to reduced seizure thresholds, behavioral abnormalities and SUDEP in various animal models of epilepsy. The levels of 3-hydroxy-4-methoxyphenylglycol, 5-hydroxyindolacetic acid and homovanillic acid in CSF samples drawn from the cisterna magna were analyzed using high-performance liquid chromatography. These levels were compared between baboons with seizures (SZ), craniofacial trauma (CFT) and asymptomatic, control (CTL) baboons, between baboons with abnormal and normal EEG studies. We hypothesized that the CSF levels of major monoaminergic metabolites (i.e., dopamine, serotonin and norepinephrine) associate with the baboons' electroclinical status and thus can be used as clinical biomarkers applicable to seizures/epilepsy. However, despite apparent differences in metabolite levels between the groups, usually lower in SZ and CFT baboons and in baboons with abnormal EEG studies, we did not find any statistically significant differences using a logistic regression analysis. Significant correlations between the metabolite levels, especially between 5-HIAA and HVA, were preserved in all electroclinical groups. While we were not able to demonstrate significant differences in monoamine metabolites in relation to seizures or EEG markers of epilepsy, we cannot exclude the monoaminergic system as a potential source of pathogenesis in epilepsy and SUDEP. A prospective study evaluating serial CSF monoamine levels in baboons with recently witnessed seizures, and evaluation of abnormal expression and function of monoaminergic receptors and transporters within epilepsy-related brain regions, may impact the electroclinical status.

Projection target-specific action of nicotine in the caudal nucleus of the solitary tract.

The brainstem nucleus of the solitary tract (NTS) is the key integrating relay in the central processing of sensory information from the thoracic and from most subdiaphragmatic viscera. Modulation of neuronal excitability and synaptic activity in the NTS by nicotinic agents can have potent effects on vital physiological functions, such as feeding, digestion, respiration, and blood circulation. Caudal NTS neurons demonstrate considerable heterogeneity in projection targets, synaptic properties, and expression of nicotinic acetylcholine receptors (nAChRs). However, despite its heterogeneity, the caudal NTS may contain discrete subsets of neurons with unique projection target-specific properties. To test this hypothesis, we used in vivo fluorescent tracing and ex vivo patch-clamp electrophysiology to evaluate responsiveness to nicotine of anatomically identified caudal NTS neurons that project to the hypothalamic paraventricular nucleus (PVN) and the brainstem caudal ventrolateral medulla (CVLM). The results of this study demonstrate that responsiveness to nicotine correlates with where the neurons project. Specifically, PVN-projecting caudal NTS neurons respond to nicotine only presynaptically (i.e., via activation of presynaptic nAChRs and potentiation of synaptic release of glutamate), suggesting indirect, glutamate-dependent effects of nicotine on the PVN-projecting NTS circuitry. By contrast, CVLM-projecting caudal NTS neurons exhibit only limited presynaptic, but dominant somatodendritic, responsiveness to nicotine, suggesting that the effects of nicotine on the CVLM-projecting NTS circuitry are direct and largely glutamate independent. Understanding the relationships among function-specific brainstem/hypothalamic neuronal networks, nuclei, and individual neurons could help develop therapies targeting identifiable neuronal circuits to offset impaired autonomic homeostasis.

The therapeutic promise of positive allosteric modulation of nicotinic receptors.

In the central nervous system, deficits in cholinergic neurotransmission correlate with decreased attention and cognitive impairment, while stimulation of neuronal nicotinic acetylcholine receptors improves attention, cognitive performance and neuronal resistance to injury as well as produces robust analgesic and anti-inflammatory effects. The rational basis for the therapeutic use of orthosteric agonists and positive allosteric modulators (PAMs) of nicotinic receptors arises from the finding that functional nicotinic receptors are ubiquitously expressed in neuronal and non-neuronal tissues including brain regions highly vulnerable to traumatic and ischemic types of injury (e.g., cortex and hippocampus). Moreover, functional nicotinic receptors do not vanish in age-, disease- and trauma-related neuropathologies, but their expression and/or activation levels decline in a subunit- and brain region-specific manner. Therefore, augmenting the endogenous cholinergic tone by nicotinic agents is possible and may offset neurological impairments associated with cholinergic hypofunction. Importantly, because neuronal damage elevates extracellular levels of choline (a selective agonist of α7 nicotinic acetylcholine receptors) near the site of injury, α7-PAM-based treatments may augment pathology-activated α7-dependent auto-therapies where and when they are most needed (i.e., in the penumbra, post-injury). Thus, nicotinic-PAM-based treatments are expected to augment the endogenous cholinergic tone in a spatially and temporally restricted manner creating the potential for differential efficacy and improved safety as compared to exogenous orthosteric nicotinic agonists that activate nicotinic receptors indiscriminately. In this review, I will summarize the existing trends in therapeutic applications of nicotinic PAMs.

Evidence for the exclusive expression of functional homomeric α7 nAChRs in hypothalamic histaminergic tuberomammillary neurons in rats.

Hypothalamic histaminergic tuberomammillary (TM) neurons in rats express high densities of nicotinic acetylcholine receptors (nAChRs) whose Ca(2+) permeability, kinetic and pharmacological properties are similar to those of heterologous homomeric α7 nAChRs. However, native α7 nAChR subunits can co-assemble with ß or α5 nAChR subunits to form functional heteromeric α7-containing α7ß or α7α5 nAChRs with kinetics and pharmacology similar to those of α7 homomers. Therefore, although TM nAChRs have been used as an ex vivo model of functional α7 homomers, the molecular makeup of TM nAChRs has not been determined and the expression of functional α7-containing heteromers in TM neurons has not been excluded. To determine the profile of TM nAChR subunit transcripts, we have conducted single-cell qRT-PCR experiments using acutely dissociated TM neurons in rats. TM neurons were found to express transcripts of only principal α3, α6 and α7 nAChR subunits. Transcripts of other known mammalian neuronal subunits (α2, α4-5, α9-10, ß2-4) were not detected. In the absence of ß and α5 subunits, the expression of functional α7-containing heteromers in TM neurons is highly unlikely because principal α3, α6 and α7 nAChR subunits alone are not known to form functional heteromeric nAChRs. These results support the exclusive expression of native functional α7 homomers in rat TM neurons and introduce these neurons as a unique reliable source of native functional homomeric α7 nAChRs suitable for ex vivo and in vitro pharmacological assays in developing selective α7 nAChR agents.

The dual effect of PNU-120596 on α7 nicotinic acetylcholine receptor channels.

PNU-120596 (1-(5-chloro-2,4-dimethoxyphenyl)-3-(5-methylisoxazol-3-yl)urea), a Type-II positive allosteric modulator of α(7) nicotinic acetylcholine receptors inhibits α(7) desensitization and robustly prolongs openings of α(7) channels. However, these effects may render α(7) channels more accessible to positively charged molecules and thus, more susceptible to voltage-dependent open-channel-block-like inhibition. To test this hypothesis, choline chloride (i.e., choline), a selective endogenous α(7) agonist, and bicuculline methochloride (i.e., bicuculline), a competitive α(7) antagonist, were used as membrane voltage-sensitive probes in whole-cell voltage-clamp recordings from hippocampal CA1 interneurons in acute brain slices in the absence and presence of PNU-120596. PNU-120596 enhanced voltage-dependent inhibition of α(7) responses by bicuculline and choline. In the presence of PNU-120596, α(7) channels favored a burst-like kinetic modality in the presence, but not absence of bicuculline and bursts of α(7) openings were voltage-dependent. These results suggest that PNU-120596 alters the pharmacology of α(7) channels by making these channels more susceptible to voltage-dependent inhibitory interactions with positively charged drugs at concentrations that do not potently inhibit α(7) channels without PNU-120596. This inhibition imitates α(7) nicotinic receptor desensitization and compromises the potentiating anti-desensitization effects of PNU-120596 on α(7) nicotinic receptors. This unexpected dual action of PNU-120596, and possibly other Type-II positive allosteric modulators of α(7) nicotinic receptors, may lead to unanticipated α(7) channel-drug interactions and misinterpretation of α(7) single-channel data.