ABSTRACT
Focused ultrasound (FUS) is a powerful tool for noninvasive modulation of deep brain activity with promising therapeutic potential for refractory epilepsy; however, tools for examining FUS effects on specific cell types within the deep brain do not yet exist. Consequently, how cell types within heterogeneous networks can be modulated and whether parameters can be identified to bias these networks in the context of complex behaviors remains unknown. To address this, we developed a fiber Photometry Coupled focused Ultrasound System (PhoCUS) for simultaneously monitoring FUS effects on neural activity of subcortical genetically targeted cell types in freely behaving animals. We identified a parameter set that selectively increases activity of parvalbumin interneurons while suppressing excitatory neurons in the hippocampus. A net inhibitory effect localized to the hippocampus was further confirmed through whole brain metabolic imaging. Finally, these inhibitory selective parameters achieved significant spike suppression in the kainate model of chronic temporal lobe epilepsy, opening the door for future noninvasive therapies.
Subject(s)
Epilepsy, Temporal Lobe , Epilepsy , Animals , Epilepsy/therapy , Brain/diagnostic imaging , Brain/physiology , Ultrasonography , Hippocampus/diagnostic imagingABSTRACT
Cessation of cannabinoid use in humans often leads to a withdrawal state that includes sleep disruption. Despite important health implications, little is known about how cannabinoid abstention affects sleep architecture, in part because spontaneous cannabinoid withdrawal is difficult to model in animals. In concurrent work we report that repeated administration of the high-efficacy cannabinoid 1 (CB1) receptor agonist AM2389 to mice for 5 days led to heightened locomotor activity and paw tremor following treatment discontinuation, potentially indicative of spontaneous cannabinoid withdrawal. Here, we performed parallel studies to examine effects on sleep. Using implantable electroencephalography (EEG) and electromyography (EMG) telemetry we examined sleep and neurophysiological measures before, during, and after 5 days of twice-daily AM2389 injections. We report that AM2389 produces decreases in locomotor activity that wane with repeated treatment, whereas discontinuation produces rebound increases in activity that persist for several days. Likewise, AM2389 initially produces profound increases in slow-wave sleep (SWS) and decreases in rapid eye movement (REM) sleep, as well as consolidation of sleep. By the third AM2389 treatment, this pattern transitions to decreases in SWS and total time sleeping. This pattern persists following AM2389 discontinuation and is accompanied by emergence of sleep fragmentation. Double-labeling immunohistochemistry for hypocretin/orexin (a sleep-regulating peptide) and c-Fos (a neuronal activity marker) in lateral hypothalamus revealed decreases in c-Fos/orexin+ cells following acute AM2389 and increases following discontinuation, aligning with the sleep changes. These findings indicate that AM2389 profoundly alters sleep in mice and suggest that sleep disruption following treatment cessation reflects spontaneous cannabinoid withdrawal.
Subject(s)
Cannabinoids , Animals , Cannabinoid Receptor Agonists/pharmacology , Cannabinoids/pharmacology , Electroencephalography , Male , Mice , Orexins , Sleep , Sleep, REM/physiologyABSTRACT
STUDY OBJECTIVE: Throughout the coronavirus disease 2019 pandemic, many emergency departments have been using passive protective enclosures ("intubation boxes") during intubation. The effectiveness of these enclosures remains uncertain. We sought to quantify their ability to contain aerosols using industry standard test protocols. METHODS: We tested a commercially available passive protective enclosure representing the most common design and compared this with a modified enclosure that incorporated a vacuum system for active air filtration during simulated intubations and negative-pressure isolation. We evaluated the enclosures by using the same 3 tests air filtration experts use to certify class I biosafety cabinets: visual smoke pattern analysis using neutrally buoyant smoke, aerosol leak testing using a test aerosol that mimics the size of virus-containing particulates, and air velocity measurements. RESULTS: Qualitative evaluation revealed smoke escaping from all passive enclosure openings. Aerosol leak testing demonstrated elevated particle concentrations outside the enclosure during simulated intubations. In contrast, vacuum-filter-equipped enclosures fully contained the visible smoke and test aerosol to standards consistent with class I biosafety cabinet certification. CONCLUSION: Passive enclosures for intubation failed to contain aerosols, but the addition of a vacuum and active air filtration reduced aerosol spread during simulated intubation and patient isolation.
Subject(s)
COVID-19/prevention & control , Infection Control/instrumentation , Intubation, Intratracheal/instrumentation , Pneumonia, Viral/prevention & control , Aerosols , COVID-19/transmission , Cross Infection/prevention & control , Equipment Design , Filtration/instrumentation , Humans , Infectious Disease Transmission, Patient-to-Professional/prevention & control , Manikins , Pandemics , Pneumonia, Viral/transmission , Pneumonia, Viral/virology , SARS-CoV-2 , VacuumABSTRACT
The lateral habenula (LHb) is involved in reward and aversion and is reciprocally connected with dopamine (DA)-containing brain regions, including the ventral tegmental area (VTA). We used a multidisciplinary approach to examine the properties of DA afferents to the LHb in the rat. We find that >90% of VTA tyrosine hydroxylase (TH) neurons projecting to the LHb lack vesicular monoamine transporter 2 (VMAT2) mRNA, and there is little coexpression of TH and VMAT2 protein in this mesohabenular pathway. Consistent with this, electrical stimulation of LHb did not evoke DA-like signals, assessed with fast-scan cyclic voltammetry. However, electrophysiological currents that were inhibited by L741,742, a DA-D4-receptor antagonist, were observed in LHb neurons when DA uptake or degradation was blocked. To prevent DA activation of D4 receptors, we repeated this experiment in LHb slices from DA-depleted rats. However, this did not disrupt D4 receptor activation initiated by the dopamine transporter inhibitor, GBR12935. As the LHb is also targeted by noradrenergic afferents, we examined whether GBR12935 activation of DA-D4 receptors occurred in slices depleted of norepinephrine (NE). Unlike DA, NE depletion prevented the activation of DA-D4 receptors. Moreover, direct application of NE elicited currents in LHb neurons that were blocked by L741,742, and GBR12935 was found to be a more effective blocker of NE uptake than the NE-selective transport inhibitor nisoxetine. These findings demonstrate that NE is released in the rat LHb under basal conditions and that it activates DA-D4 receptors. Therefore, NE may be an important regulator of LHb function.
Subject(s)
Habenula/metabolism , Norepinephrine/pharmacology , Receptors, Dopamine D4/metabolism , Animals , Dopamine/metabolism , Dopamine/pharmacology , Dopamine Agonists/pharmacology , Dopamine Antagonists/pharmacology , Dopamine Plasma Membrane Transport Proteins/antagonists & inhibitors , Dopaminergic Neurons/metabolism , Dopaminergic Neurons/physiology , Habenula/cytology , Habenula/physiology , Isoxazoles/pharmacology , Male , Norepinephrine/metabolism , Piperazines/pharmacology , Piperidines/pharmacology , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Receptors, Dopamine D4/antagonists & inhibitors , Ventral Tegmental Area/cytology , Ventral Tegmental Area/metabolism , Ventral Tegmental Area/physiology , Vesicular Monoamine Transport Proteins/genetics , Vesicular Monoamine Transport Proteins/metabolismABSTRACT
Glutamatergic lateral habenula (LHb) output communicates negative motivational valence to ventral tegmental area (VTA) dopamine (DA) neurons via activation of the rostromedial tegmental nucleus (RMTg). However, the LHb also receives a poorly understood DA input from the VTA, which we hypothesized constitutes an important feedback loop regulating DA responses to stimuli. Using whole-cell electrophysiology in rat brain slices, we find that DA initiates a depolarizing inward current (I(DAi)) and increases spontaneous firing in 32% of LHb neurons. I(DAi) was also observed upon application of amphetamine or the DA uptake blockers cocaine or GBR12935, indicating involvement of endogenous DA. I(DAi) was blocked by D4 receptor (D4R) antagonists (L745,870 or L741,742), and mimicked by a selective D4R agonist (A412997). I(DAi) was associated with increased whole-cell conductance and was blocked by Cs+ or a selective blocker of hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channel, ZD7288. I(DAi) was also associated with a depolarizing shift in half-activation voltage for the hyperpolarization-activated cation current (Ih) mediated by HCN channels. Recordings from LHb neurons containing fluorescent retrograde tracers revealed that I(DAi) was observed only in cells projecting to the RMTg and not the VTA. In parallel with direct depolarization, DA also strongly increased synaptic glutamate release and reduced synaptic GABA release onto LHb cells. These results demonstrate that DA can excite glutamatergic LHb output to RMTg via multiple cellular mechanisms. Since the RMTg strongly inhibits midbrain DA neurons, activation of LHb output to RMTg by DA represents a negative feedback loop that may dampen DA neuron output following activation.
Subject(s)
Action Potentials , Habenula/physiology , Neurons/physiology , Receptors, Dopamine D4/metabolism , Amphetamine/pharmacology , Animals , Cesium/pharmacology , Cocaine/pharmacology , Dopamine/pharmacology , Dopamine Agonists/pharmacology , Dopamine Antagonists/pharmacology , Dopamine Uptake Inhibitors/pharmacology , Feedback, Physiological , Glutamic Acid/metabolism , Habenula/cytology , Habenula/metabolism , Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels/antagonists & inhibitors , Male , Neurons/drug effects , Neurons/metabolism , Pedunculopontine Tegmental Nucleus/cytology , Pedunculopontine Tegmental Nucleus/physiology , Piperazines/pharmacology , Rats , Rats, Sprague-Dawley , Receptors, Dopamine D4/agonists , Receptors, Dopamine D4/antagonists & inhibitors , Synaptic Transmission , Ventral Tegmental Area/cytology , Ventral Tegmental Area/physiology , gamma-Aminobutyric Acid/metabolismABSTRACT
Many strong rewards, including abused drugs, also produce aversive effects that are poorly understood. For example, cocaine can produce aversive conditioning after its rewarding effects have dissipated, consistent with opponent process theory, but the neural mechanisms involved are not well known. Using electrophysiological recordings in awake rats, we found that some neurons in the lateral habenula (LHb), where activation produces aversive conditioning, exhibited biphasic responses to single doses of intravenous cocaine, with an initial inhibition followed by delayed excitation paralleling cocaine's shift from rewarding to aversive. Recordings in LHb slice preparations revealed similar cocaine-induced biphasic responses and further demonstrated that biphasic responses were mimicked by dopamine, that the inhibitory phase depended on dopamine D2-like receptors, and that the delayed excitation persisted after drug washout for prolonged durations consistent with findings in vivo. c-Fos experiments further showed that cocaine-activated LHb neurons preferentially projected to and activated neurons in the rostromedial tegmental nucleus (RMTg), a recently identified target of LHb axons that is activated by negative motivational stimuli and inhibits dopamine neurons. Finally, pharmacological excitation of the RMTg produced conditioned place aversion, whereas cocaine-induced avoidance behaviors in a runway operant paradigm were abolished by lesions of LHb efferents, lesions of the RMTg, or by optogenetic inactivation of the RMTg selectively during the period when LHb neurons are activated by cocaine. Together, these results indicate that LHb/RMTg pathways contribute critically to cocaine-induced avoidance behaviors, while also participating in reciprocally inhibitory interactions with dopamine neurons.
Subject(s)
Avoidance Learning/drug effects , Cocaine/administration & dosage , Conditioning, Operant/drug effects , Dopamine , Habenula/drug effects , Mesencephalon/drug effects , Animals , Avoidance Learning/physiology , Conditioning, Operant/physiology , Dopamine/physiology , Habenula/physiology , Injections, Intravenous , Male , Mesencephalon/physiology , Neural Pathways/drug effects , Neural Pathways/physiology , Rats , Rats, Sprague-Dawley , Rats, WistarABSTRACT
Neurotechnologies and genetic tools for dissecting neural circuit functions have advanced rapidly over the past decade, although the development of complementary pharmacological method-ologies has comparatively lagged. Understanding the precise pharmacological mechanisms of neuroactive compounds is critical for advancing basic neurobiology and neuropharmacology, as well as for developing more effective treatments for neurological and neuropsychiatric disorders. However, integrating modern tools for assessing neural activity in large-scale neural networks with spatially localized drug delivery remains a major challenge. Here, we present a dual microfluidic-photometry platform that enables simultaneous intracranial drug delivery with neural dynamics monitoring in the rodent brain. The integrated platform combines a wireless, battery-free, miniaturized fluidic microsystem with optical probes, allowing for spatially and temporally specific drug delivery while recording activity-dependent fluorescence using genetically encoded calcium indicators (GECIs), neurotransmitter sensors GRAB NE and GRAB DA , and neuropeptide sensors. We demonstrate the performance this platform for investigating neuropharmacological mechanisms in vivo and characterize its efficacy in probing precise mechanistic actions of neuroactive compounds across several rapidly evolving neuroscience domains.
ABSTRACT
Comprehensive, continuous quantitative monitoring of intricately orchestrated physiological processes and behavioral states in living organisms can yield essential data for elucidating the function of neural circuits under healthy and diseased conditions, for defining the effects of potential drugs and treatments, and for tracking disease progression and recovery. Here, we report a wireless, battery-free implantable device and a set of associated algorithms that enable continuous, multiparametric physio-behavioral monitoring in freely behaving small animals and interacting groups. Through advanced analytics approaches applied to mechano-acoustic signals of diverse body processes, the device yields heart rate, respiratory rate, physical activity, temperature, and behavioral states. Demonstrations in pharmacological, locomotor, and acute and social stress tests and in optogenetic studies offer unique insights into the coordination of physio-behavioral characteristics associated with healthy and perturbed states. This technology has broad utility in neuroscience, physiology, behavior, and other areas that rely on studies of freely moving, small animal models.
Subject(s)
Behavior, Animal , Optogenetics , Wireless Technology , Animals , Behavior, Animal/physiology , Optogenetics/methods , Mice , Heart Rate/physiology , Male , Prostheses and Implants , Respiratory Rate/physiology , Monitoring, Physiologic/methods , Monitoring, Physiologic/instrumentation , AlgorithmsABSTRACT
Fully implantable wireless systems for the recording and modulation of neural circuits that do not require physical tethers or batteries allow for studies that demand the use of unconstrained and freely behaving animals in isolation or in social groups. Moreover, feedback-control algorithms that can be executed within such devices without the need for remote computing eliminate virtual tethers and any associated latencies. Here we report a wireless and battery-less technology of this type, implanted subdermally along the back of freely moving small animals, for the autonomous recording of electroencephalograms, electromyograms and body temperature, and for closed-loop neuromodulation via optogenetics and pharmacology. The device incorporates a system-on-a-chip with Bluetooth Low Energy for data transmission and a compressed deep-learning module for autonomous operation, that offers neurorecording capabilities matching those of gold-standard wired systems. We also show the use of the implant in studies of sleep-wake regulation and for the programmable closed-loop pharmacological suppression of epileptic seizures via feedback from electroencephalography. The technology can support a broader range of applications in neuroscience and in biomedical research with small animals.
ABSTRACT
Parkinson's disease (PD) involves progressive loss of nigrostriatal dopamine (DA) neurons over an extended period of time. Mitochondrial damage may lead to PD, and neurotoxins affecting mitochondria are widely used to produce degeneration of the nigrostriatal circuitry. Deletion of the mitochondrial transcription factor A gene (Tfam) in C57BL6 mouse DA neurons leads to a slowly progressing parkinsonian phenotype in which motor impairment is first observed at ~12 wk of age. L-DOPA treatment improves motor dysfunction in these "MitoPark" mice, but this declines when DA neuron loss is more complete. To investigate early neurobiological events potentially contributing to PD, we compared the neurochemical and electrophysiological properties of the nigrostriatal circuit in behaviorally asymptomatic 6- to 8-wk-old MitoPark mice and age-matched control littermates. Release, but not uptake of DA, was impaired in MitoPark mouse striatal brain slices, and nigral DA neurons lacked characteristic pacemaker activity compared with control mice. Also, hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channel function was reduced in MitoPark DA neurons, although HCN messenger RNA was unchanged. This study demonstrates altered nigrostriatal function that precedes behavioral parkinsonian symptoms in this genetic PD model. A full understanding of these presymptomatic cellular properties may lead to more effective early treatments of PD.
Subject(s)
Cyclic Nucleotide-Gated Cation Channels/physiology , DNA-Binding Proteins/genetics , Mitochondrial Proteins/genetics , Neurons/physiology , Parkinson Disease/physiopathology , Transcription Factors/genetics , Animals , Corpus Striatum , Disease Models, Animal , Dopamine/metabolism , Female , Male , Mice , Mice, Inbred C57BL , Neurons/metabolism , Parkinson Disease/genetics , Substantia NigraABSTRACT
Wireless battery-free optogenetic devices enable behavioral neuroscience studies in groups of animals with minimal interference to natural behavior. Real-time independent control of optogenetic stimulation through near-field communication dramatically expands the realm of applications of these devices in broad contexts of neuroscience research. Dissemination of these tools with advanced functionalities to the neuroscience community requires protocols for device manufacturing and experimental implementation. This protocol describes detailed procedures for fabrication, encapsulation and implantation of recently developed advanced wireless devices in head- and back-mounted forms. In addition, procedures for standard implementation of experimental systems in mice are provided. This protocol aims to facilitate the application of wireless optogenetic devices in advanced optogenetic experiments involving groups of freely moving rodents and complex environmental designs. The entire protocol lasts ~3-5 weeks.
Subject(s)
Neurosciences , Optogenetics , Animals , Mice , Optogenetics/methods , Wireless TechnologyABSTRACT
Ventral tegmental area (VTA) dopamine (DA) neurons play a pivotal role in processing reward-related information and are involved in drug addiction and mental illness in humans. Information is conveyed to the VTA in large part by glutamatergic afferents that arise in various brain nuclei, including the pedunculopontine nucleus (PPN). Using a unique rat brain slice preparation, we found that PPN stimulation activates afferents targeting GluR2-containing AMPA receptors (AMPAR) on VTA DA neurons, and these afferents did not exhibit long-term depression (LTD). In contrast, activation of glutamate afferents onto the same DA neurons via stimulation within the VTA evoked EPSCs mediated by GluR2-lacking AMPARs that demonstrated LTD or EPSCs mediated by GluR2-containing AMPA receptors that did not express LTD. Twenty-four hours after single cocaine injections to rats, GluR2-lacking AMPARs were increased at both PPN and local VTA projections, and this permitted LTD expression in both pathways. Single injections with the main psychoactive ingredient of marijuana, Delta(9)-tetrahydrocannabinol (Delta(9)-THC), increased GluR2-lacking AMPA receptors and permitted LTD in only the PPN pathway, and these effects were prevented by in vivo pretreatment with the cannabinoid CB1 receptor antagonist AM251. These results demonstrate that cocaine more globally increases GluR2-lacking AMPA receptors at all glutamate synapses on VTA dopamine neurons, whereas Delta(9)-THC selectively increased GluR2-lacking AMPA receptors at subcortical PPN synapses. This suggests that different abused drugs may exert influence over distinct sets of glutamatergic afferents to VTA DA neurons which may be associated with different reinforcing or addictive properties of these drugs.
Subject(s)
Cocaine/pharmacology , Dopamine/metabolism , Dronabinol/pharmacology , Mesencephalon/metabolism , Neuronal Plasticity/drug effects , Psychotropic Drugs/pharmacology , Receptors, AMPA/metabolism , Synapses/metabolism , Animals , Dopamine Uptake Inhibitors/pharmacology , Excitatory Postsynaptic Potentials/drug effects , Illicit Drugs/pharmacology , Immunohistochemistry , Injections, Intraperitoneal , Long-Term Synaptic Depression/drug effects , Male , Mesencephalon/cytology , Mesencephalon/drug effects , Neurons, Afferent/drug effects , Neurons, Afferent/metabolism , Organ Culture Techniques , Pedunculopontine Tegmental Nucleus/metabolism , Rats , Rats, Sprague-Dawley , Synapses/drug effects , Ventral Tegmental Area/metabolismABSTRACT
Advanced technologies for controlled delivery of light to targeted locations in biological tissues are essential to neuroscience research that applies optogenetics in animal models. Fully implantable, miniaturized devices with wireless control and power-harvesting strategies offer an appealing set of attributes in this context, particularly for studies that are incompatible with conventional fiber-optic approaches or battery-powered head stages. Limited programmable control and narrow options in illumination profiles constrain the use of existing devices. The results reported here overcome these drawbacks via two platforms, both with real-time user programmability over multiple independent light sources, in head-mounted and back-mounted designs. Engineering studies of the optoelectronic and thermal properties of these systems define their capabilities and key design considerations. Neuroscience applications demonstrate that induction of interbrain neuronal synchrony in the medial prefrontal cortex shapes social interaction within groups of mice, highlighting the power of real-time subject-specific programmability of the wireless optogenetic platforms introduced here.
Subject(s)
Optogenetics/instrumentation , Social Behavior , Wireless Technology/instrumentation , Animals , MiceABSTRACT
The military lifestyle often includes continuous operations whether in training or deployed environments. These stressful environments present unique challenges for service members attempting to achieve consolidated, restorative sleep. The significant mental and physical derangements caused by degraded metabolic, cardiovascular, skeletomuscular, and cognitive health often result from insufficient sleep and/or circadian misalignment. Insufficient sleep and resulting fatigue compromises personal safety, mission success, and even national security. In the long-term, chronic insufficient sleep and circadian rhythm disorders have been associated with other sleep disorders (e.g., insomnia, obstructive sleep apnea, and parasomnias). Other physiologic and psychologic diagnoses such as post-traumatic stress disorder, cardiovascular disease, and dementia have also been associated with chronic, insufficient sleep. Increased co-morbidity and mortality are compounded by traumatic brain injury resulting from blunt trauma, blast exposure, and highly physically demanding tasks under load. We present the current state of science in human and animal models specific to service members during- and post-military career. We focus on mission requirements of night shift work, sustained operations, and rapid re-entrainment to time zones. We then propose targeted pharmacological and non-pharmacological countermeasures to optimize performance that are mission- and symptom-specific. We recognize a critical gap in research involving service members, but provide tailored interventions for military health care providers based on the large body of research in health care and public service workers.
Subject(s)
Military Personnel/psychology , Sleep Disorders, Circadian Rhythm/physiopathology , Sleep Disorders, Circadian Rhythm/psychology , Sleep/physiology , Animals , Humans , Sleep Disorders, Circadian Rhythm/epidemiology , Sleep Wake Disorders/epidemiology , Sleep Wake Disorders/physiopathology , Sleep Wake Disorders/psychology , Stress Disorders, Post-Traumatic/epidemiology , Stress Disorders, Post-Traumatic/physiopathology , Stress Disorders, Post-Traumatic/psychology , United States/epidemiologyABSTRACT
Anatomical studies indicate that synaptic inputs from many cortical and subcortical structures converge on neurons of the ventral tegmental area (VTA). Although in vitro electrophysiological studies have examined synaptic inputs to dopamine (DA) and non-DA neurons in the VTA, they have largely relied upon local electrical stimulation to activate these synapses. This provides little information regarding the distinct properties of synapses originating from different brain areas. Using whole-cell recordings in parasagittal rat brain slices that preserved subcortical axons from the pedunculopontine nucleus (PPN) to the VTA, we compared these synapses with those activated by intra-VTA stimulation. PPN-evoked currents demonstrated longer latencies than intra-VTA-evoked currents, and both VTA and PPN responses were mediated by GABA(A) and AMPA receptors. However, unlike VTA-evoked currents, PPN currents were exclusively mediated by glutamate in 25-40% of the VTA neurons. Consistent with a cholinergic projection from the PPN to the VTA, nicotinic acetylcholine receptors (nAChR) were activated by endogenous acetylcholine released during PPN, but not VTA, stimulation. This was seen as a reduction of PPN-evoked, and not VTA-evoked, synaptic currents by the alpha7-nAChR antagonist methyllycaconitine (MLA) and the agonist nicotine. The beta2-nAChR subunit antagonist dihydro-beta-erythroidine had no effect on VTA- or PPN-evoked synaptic currents. The effects of MLA on PPN-evoked currents were unchanged by the GABA(A) receptor blocker picrotoxin, indicating that alpha7-nAChRs presynaptically modulated glutamate and not GABA release. These differences in physiological and pharmacological properties demonstrate that ascending PPN and presumed descending inputs to VTA utilize distinct mechanisms to differentially modulate neuronal activity and encode cortical and subcortical information.
Subject(s)
Afferent Pathways/physiology , Pedunculopontine Tegmental Nucleus/physiology , Synapses/physiology , Synaptic Transmission/physiology , Ventral Tegmental Area/physiology , Aconitine/analogs & derivatives , Aconitine/pharmacology , Afferent Pathways/drug effects , Animals , Bicuculline/pharmacology , Dihydro-beta-Erythroidine/pharmacology , Electric Stimulation , Excitatory Amino Acid Antagonists/pharmacology , GABA Antagonists/pharmacology , In Vitro Techniques , Nicotine/pharmacology , Nicotinic Antagonists/pharmacology , Patch-Clamp Techniques , Pedunculopontine Tegmental Nucleus/drug effects , Picrotoxin/pharmacology , Quinoxalines/pharmacology , Rats , Rats, Sprague-Dawley , Reaction Time/drug effects , Reaction Time/physiology , Receptors, Nicotinic/physiology , Strontium/pharmacology , Synapses/drug effects , Synaptic Transmission/drug effects , Ventral Tegmental Area/drug effects , alpha7 Nicotinic Acetylcholine ReceptorABSTRACT
The rostromedial tegmental nucleus (RMTg), also known as the tail of the ventral tegmental area (tVTA), is a GABAergic structure identified in 2009 that receives strong inputs from the lateral habenula and other sources, sends dense inhibitory projections to midbrain dopamine (DA) neurons, and plays increasingly recognized roles in aversive learning, addiction, and other motivated behaviors. In general, little is known about the genetic identity of these neurons. However, recent work has identified the transcription factor FoxP1 as enhanced in the mouse RMTg (Lahti et al. in Development 143(3):516-529, 2016). Hence, in the current study, we used RNA sequencing to identify genes significantly enhanced in the rat RMTg as compared to adjacent VTA, and then examined the detailed distribution of two genes in particular, prepronociceptin (Pnoc) and FoxP1. In rats and mice, both Pnoc and FoxP1 were expressed at high levels in the RMTg and colocalized strongly with previously established RMTg markers. FoxP1 was particularly selective for RMTg neurons, as it was absent in most adjacent brain regions. We used these gene expression patterns to refine the anatomic characterization of RMTg in rats, extend this characterization to mice, and show that optogenetic manipulation of RMTg in mice bidirectionally modulates real-time place preference. Hence, RMTg neurons in both rats and mice exhibit distinct genetic profiles that correlate with their distinct connectivity and function.
Subject(s)
Forkhead Transcription Factors/metabolism , Neurons/metabolism , Protein Precursors/metabolism , Receptors, Opioid/metabolism , Repressor Proteins/metabolism , Ventral Tegmental Area/metabolism , Animals , Behavior, Animal , Forkhead Transcription Factors/genetics , Gene Expression Regulation , Male , Mice, Transgenic , Motor Activity , Neural Pathways/metabolism , Optogenetics , Protein Precursors/genetics , Proto-Oncogene Proteins c-fos/metabolism , Rats, Sprague-Dawley , Receptors, Opioid/genetics , Repressor Proteins/genetics , Time Factors , Ventral Tegmental Area/cytologyABSTRACT
The pedunculopontine nucleus (PPN), the cholinergic arm of the reticular activating system (RAS), is known to modulate waking and rapid eye movement (REM) sleep. REM sleep decreases between 10 and 30 days postnatally in the rat, with the majority occurring between 12 and 21 days. We investigated the possibility that changes in the cholinergic, muscarinic and/or nicotinic, input to PPN neurons could explain at least part of the developmental decrease in REM sleep. We recorded intracellularly from PPN neurons in 12-21 day rat brainstem slices maintained in artificial cerebrospinal fluid (aCSF) and found that application of the nicotinic agonist 1,1-dimethyl-4-phenyl-piperazinium iodide (DMPP) depolarized PPN neurons early in development, then hyperpolarized PPN neurons by day 21. Most of the effects of DMPP persisted following application of the sodium channel blocker tetrodotoxin (TTX), and in the presence of glutamatergic, serotonergic, noradrenergic and GABAergic antagonists, but were blocked by the nicotinic antagonist mecamylamine (MEC). The mixed muscarinic agonist carbachol (CAR) hyperpolarized all type II (A current) PPN cells and depolarized all type I (low threshold spike-LTS current) and type III (A+LTS current) PPN cells, but did not change effects during the period known for the developmental decrease in REM sleep. The effects of CAR persisted in the presence of TTX but were mostly blocked by the muscarinic antagonist atropine (ATR), and the remainder by MEC. We conclude that, while the nicotinic inputs to the PPN may help modulate the developmental decrease in REM sleep, the muscarinic inputs appear to modulate different types of cells differentially.