Cocaine-induced endocannabinoid signaling mediated by sigma-1 receptors and extracellular vesicle secretion.

Cocaine is an addictive drug that acts in brain reward areas. Recent evidence suggests that cocaine stimulates synthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG) in midbrain, increasing dopamine neuron activity via disinhibition. Although a mechanism for cocaine-stimulated 2-AG synthesis is known, our understanding of 2-AG release is limited. In NG108 cells and mouse midbrain tissue, we find that 2-AG is localized in non-synaptic extracellular vesicles (EVs) that are secreted in the presence of cocaine via interaction with the chaperone protein sigma-1 receptor (Sig-1R). The release of EVs occurs when cocaine causes dissociation of the Sig-1R from ADP-ribosylation factor (ARF6), a G-protein regulating EV trafficking, leading to activation of myosin light chain kinase (MLCK). Blockade of Sig-1R function, or inhibition of ARF6 or MLCK also prevented cocaine-induced EV release and cocaine-stimulated 2-AG-modulation of inhibitory synapses in DA neurons. Our results implicate the Sig-1R-ARF6 complex in control of EV release and demonstrate that cocaine-mediated 2-AG release can occur via EVs.

Cocaine-Induced Endocannabinoid Mobilization in the Ventral Tegmental Area.

Cocaine is a highly addictive drug that acts upon the brain's reward circuitry via the inhibition of monoamine uptake. Endogenous cannabinoids (eCB) are lipid molecules released from midbrain dopamine (DA) neurons that modulate cocaine's effects through poorly understood mechanisms. We find that cocaine stimulates release of the eCB, 2-arachidonoylglycerol (2-AG), in the rat ventral midbrain to suppress GABAergic inhibition of DA neurons, through activation of presynaptic cannabinoid CB1 receptors. Cocaine mobilizes 2-AG via inhibition of norepinephrine uptake and promotion of a cooperative interaction between Gq/11-coupled type-1 metabotropic glutamate and α1-adrenergic receptors to stimulate internal calcium stores and activate phospholipase C. The disinhibition of DA neurons by cocaine-mobilized 2-AG is also functionally relevant because it augments DA release in the nucleus accumbens in vivo. Our results identify a mechanism through which the eCB system can regulate the rewarding and addictive properties of cocaine.

FPGA implementation of principal component regression (PCR) for real-time differentiation of dopamine from interferents.

This paper reports on field-programmable gate array (FPGA) implementation of a digital signal processing (DSP) unit for real-time processing of neurochemical data obtained by fast-scan cyclic voltammetry (FSCV) at a carbonfiber microelectrode (CFM). The DSP unit comprises a decimation filter and two embedded processors to process the FSCV data obtained by an oversampling recording front-end and differentiate the target analyte from interferents in real time with a chemometrics algorithm using principal component regression (PCR). Interfaced with an integrated, FSCV-sensing front-end, the DSP unit successfully resolves the dopamine response from that of pH change and background-current drift, two common dopamine interferents, in flow injection analysis involving bolus injection of mixed solutions, as well as in biological tests involving electrically evoked, transient dopamine release in the forebrain of an anesthetized rat.

Real-time processing of fast-scan cyclic voltammetry (FSCV) data using a field-programmable gate array (FPGA).

This paper reports the hardware implementation of a digital signal processing (DSP) unit for real-time processing of data obtained by fast-scan cyclic voltammetry (FSCV) at a carbon-fiber microelectrode (CFM), an electrochemical transduction technique for high-resolution monitoring of brain neurochemistry. Implemented on a field-programmable gate array (FPGA), the DSP unit comprises a decimation filter and an embedded processor to process the oversampled FSCV data and obtain in real time a temporal profile of concentration variation along with a chemical signature to identify the target neurotransmitter. Interfaced with an integrated, FSCV-sensing front-end, the DSP unit can successfully process FSCV data obtained by bolus injection of dopamine in a flow cell as well as electrically evoked, transient dopamine release in the dorsal striatum of an anesthetized rat.

Amphetamine augments action potential-dependent dopaminergic signaling in the striatum in vivo.

Amphetamine (AMPH) is thought to disrupt normal patterns of action potential-dependent dopaminergic signaling by depleting dopamine (DA) vesicular stores and promoting non-exocytotic DA efflux. Voltammetry in brain slices concurrently demonstrates these key drug effects, along with competitive inhibition of neuronal DA uptake. Here, we perform comparable kinetic and voltammetric analyses in vivo to determine whether AMPH acts qualitatively and quantitatively similar in the intact brain. Fast-scan cyclic voltammetry measured extracellular DA in dorsal and ventral striata of urethane-anesthetized rats. Electrically evoked recordings were analyzed to determine K(m) and V(max) for DA uptake and vesicular DA release, while background voltammetric current indexed basal DA concentration. AMPH (0.5, 3, and 10 mg/kg i.p.) robustly increased evoked DA responses in both striatal subregions. The predominant contributor to these elevated levels was competitive uptake inhibition, as exocytotic release was unchanged in the ventral striatum and only modestly decreased in the dorsal striatum. Increases in basal DA levels were not detected. These results are consistent with AMPH augmenting action potential-dependent dopaminergic signaling in vivo across a wide, behaviorally relevant dose range. Future work should be directed at possible causes for the distinct in vitro and in vivo pharmacology of AMPH.

A miniaturized device for wireless FSCV monitoring of dopamine in an ambulatory subject.

This paper reports on a miniaturized device for wireless monitoring of extracellular dopamine levels in the brain of an ambulatory rat using fast-scan cyclic voltammetry at a carbon-fiber microelectrode. The device comprises integrated circuitry for neurochemical recording fabricated in 0.5-microm double-poly triple-metal CMOS technology, which is assembled and packaged on a miniature rigid-flex substrate together with a few external components for supply generation, biasing, and chip programming. The device operates from a single 3-V battery, weighs 2.3 g (including the battery), and upon implantation successfully captures the effects of the psychostimulant amphetamine on electrically and non-electrically evoked dopamine neurotransmission in the caudateputamen region of an ambulatory rat's forebrain.

Teaching brain-behavior relations economically with stimulus equivalence technology.

Instructional interventions based on stimulus equivalence provide learners with the opportunity to acquire skills that are not directly taught, thereby improving the efficiency of instructional efforts. The present report describes a study in which equivalence-based instruction was used to teach college students facts regarding brain anatomy and function. The instruction involved creating two classes of stimuli that students understood as being related. Because the two classes shared a common member, they spontaneously merged, thereby increasing the yield of emergent relations. Overall, students mastered more than twice as many facts as were explicitly taught, thus demonstrating the potential of equivalence-based instruction to reduce the amount of student investment that is required to master advanced academic topics.

Wireless Instantaneous Neurotransmitter Concentration Sensing System (WINCS) for intraoperative neurochemical monitoring.

The Wireless Instantaneous Neurotransmitter Concentration Sensing System (WINCS) measures extracellular neurotransmitter concentration in vivo and displays the data graphically in nearly real time. WINCS implements two electroanalytical methods, fast-scan cyclic voltammetry (FSCV) and fixed-potential amperometry (FPA), to measure neurotransmitter concentrations at an electrochemical sensor, typically a carbon-fiber microelectrode. WINCS comprises a battery-powered patient module and a custom software application (WINCSware) running on a nearby personal computer. The patient module impresses upon the electrochemical sensor either a constant potential (for FPA) or a time-varying waveform (for FSCV). A transimpedance amplifier converts the resulting current to a signal that is digitized and transmitted to the base station via a Bluetooth radio link. WINCSware controls the operational parameters for FPA or FSCV, and records the transmitted data stream. Filtered data is displayed in various formats, including a background-subtracted plot of sequential FSCV scans - a representation that enables users to distinguish the signatures of various analytes with considerable specificity. Dopamine, glutamate, adenosine and serotonin were selected as analytes for test trials. Proof-of-principle tests included in vitro flow-injection measurements and in vivo measurements in rat and pig. Further testing demonstrated basic functionality in a 3-Tesla MRI unit. WINCS was designed in compliance with consensus standards for medical electrical device safety, and it is anticipated that its capability for real-time intraoperative monitoring of neurotransmitter release at an implanted sensor will prove useful for advancing functional neurosurgery.

Development of the Wireless Instantaneous Neurotransmitter Concentration System for intraoperative neurochemical monitoring using fast-scan cyclic voltammetry.

OBJECT: Emerging evidence supports the hypothesis that modulation of specific central neuronal systems contributes to the clinical efficacy of deep brain stimulation (DBS) and motor cortex stimulation (MCS). Real-time monitoring of the neurochemical output of targeted regions may therefore advance functional neurosurgery by, among other goals, providing a strategy for investigation of mechanisms, identification of new candidate neurotransmitters, and chemically guided placement of the stimulating electrode. The authors report the development of a device called the Wireless Instantaneous Neurotransmitter Concentration System (WINCS) for intraoperative neurochemical monitoring during functional neurosurgery. This device supports fast-scan cyclic voltammetry (FSCV) at a carbon-fiber microelectrode (CFM) for real-time, spatially and chemically resolved neurotransmitter measurements in the brain. METHODS: The FSCV study consisted of a triangle wave scanned between -0.4 and 1 V at a rate of 300 V/second and applied at 10 Hz. All voltages were compared with an Ag/AgCl reference electrode. The CFM was constructed by aspirating a single carbon fiber (r = 2.5 mum) into a glass capillary and pulling the capillary to a microscopic tip by using a pipette puller. The exposed carbon fiber (that is, the sensing region) extended beyond the glass insulation by approximately 100 microm. The neurotransmitter dopamine was selected as the analyte for most trials. Proof-of-principle tests included in vitro flow injection and noise analysis, and in vivo measurements in urethane-anesthetized rats by monitoring dopamine release in the striatum following high-frequency electrical stimulation of the medial forebrain bundle. Direct comparisons were made to a conventional hardwired system. RESULTS: The WINCS, designed in compliance with FDA-recognized consensus standards for medical electrical device safety, consisted of 4 modules: 1) front-end analog circuit for FSCV (that is, current-to-voltage transducer); 2) Bluetooth transceiver; 3) microprocessor; and 4) direct-current battery. A Windows-XP laptop computer running custom software and equipped with a Universal Serial Bus-connected Bluetooth transceiver served as the base station. Computer software directed wireless data acquisition at 100 kilosamples/second and remote control of FSCV operation and adjustable waveform parameters. The WINCS provided reliable, high-fidelity measurements of dopamine and other neurochemicals such as serotonin, norepinephrine, and ascorbic acid by using FSCV at CFM and by flow injection analysis. In rats, the WINCS detected subsecond striatal dopamine release at the implanted sensor during high-frequency stimulation of ascending dopaminergic fibers. Overall, in vitro and in vivo testing demonstrated comparable signals to a conventional hardwired electrochemical system for FSCV. Importantly, the WINCS reduced susceptibility to electromagnetic noise typically found in an operating room setting. CONCLUSIONS: Taken together, these results demonstrate that the WINCS is well suited for intraoperative neurochemical monitoring. It is anticipated that neurotransmitter measurements at an implanted chemical sensor will prove useful for advancing functional neurosurgery.