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NMDA Receptor-Dependent Cholinergic Modulation of Mesolimbic Dopamine Cell Bodies: Neurochemical and Behavioral Studies Marina Spanos, Xiaohu Xie, Julie Gras-Najjar, Stephanie C. White, and Leslie A. Sombers ACS Chem. Neurosci., Just Accepted Manuscript • DOI: 10.1021/acschemneuro.8b00492 • Publication Date (Web): 09 Nov 2018 Downloaded from http://pubs.acs.org on November 10, 2018
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ACS Chemical Neuroscience
NMDA Receptor-Dependent Cholinergic Modulation of Mesolimbic Dopamine Cell Bodies: Neurochemical and Behavioral Studies
Marina Spanos1, Xiaohu Xie1, Julie Gras-Najjar, Stephanie C. White, Leslie A. Sombers* 1 Authors
contributed equally.
Department of Chemistry, North Carolina State University, Raleigh, NC 27695-8204
*To whom correspondence should be addressed
[email protected] Tel. (919) 515-0320
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ABSTRACT Substance abuse disorders are devastating, costly, and difficult to treat. Identifying the neurochemical mechanisms underlying reinforcement promises to provide critical information in the development of effective treatments. Several lines of evidence suggest that striatal dopamine (DA) release serves as a teaching signal in reinforcement learning, and that shifts in DA release from the primary reward to reward-predicting stimuli play a critical role in the selfadministration of both natural and non-natural rewards. However, far less is known about the reinforcing effects of motivationally neutral sensory stimuli, or how these signals can facilitate self-administration behavior. Thus, we trained rats (n=7) to perform a visual stimulus-induced instrumental task, which involved lever pressing for activation of a stimulus light. We then microinfused vehicle (phosphate buffered saline), carbachol (acetylcholine receptor agonist), or carbachol in the presence of an N-methyl-D-aspartate (NMDA) receptor-specific drug (NMDA itself, or the antagonist, AP5) into the ventral tegmental area (VTA). This enabled us to directly evaluate how chemical modulation of dopamine cell bodies affects the instrumental behavior, as well as the nature of extracellular dopamine transients recorded in the nucleus accumbens shell (NAc shell) using fast-scan cyclic voltammetry (FSCV). Intra-VTA infusion of carbachol enhanced the magnitude and frequency of dopamine transients in the NAc shell and potentiated active lever responding without altering inactive lever responding, as compared to infusion of vehicle. Co-infusion of carbachol with AP5 abolished dopamine transients recorded in the NAc and attenuated active lever responding without altering inactive lever responding. Finally, coadministration of carbachol and NMDA into the VTA restored both lever pressing and dopaminergic signals recorded in the striatum. Together, these results suggest that acetylcholine and glutamate synergistically act at dopamine cells in the VTA to modulate VTANAc shell dopaminergic output, and this underlies motivation to lever press for a motivationally neutral visual stimulus. Keywords: Glutamate, Acetylcholine, Dopamine, Sensory stimuli, Ventral Tegmental Area 2 ACS Paragon Plus Environment
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INTRODUCTION Sensory stimuli play a critical role in the initiation of food seeking and consumption
1, 2,
as well as drug use. One example is menthol, a substance that can produce a cool feeling, and is likely to promote cigarette smoking, even for first time cigarette smokers that have not previously experienced the reinforcing effects of nicotine
3-5.
Similarly, alcoholic beverages are
generally flavored prior to consumption, as the flavor contributes to the development of alcohol preference and consumption patterns in both humans and animals 6. Thus, the role of neutral sensory stimuli in promoting drug use may be partly mediated by the reinforcing effects of the sensory stimulus itself. Multiple studies have reported that sensory stimuli can function as primary reinforcers. For instance, animal studies have shown that the primary reinforcing effects of non-conditioned neutral sensory stimuli (e.g. a stimulus light) play an important role in nicotine7-12 and methamphetamine13 self-administration experiments. However, the neural mechanisms underlying this phenomenon have not been fully investigated. Behaviorally salient stimuli are thought to initiate burst firing of dopamine (DA) neurons in the ventral tegmental area (VTA) as a key step in reward processing 14-16. These DA neurons are regulated by many inputs that contain a variety of chemical modulators ranging from small molecules to larger neuropeptides. For instance, a large excitatory input originates from two glutamatergic/cholinergic groups of mesopontine tegmental area neurons: the pedunculopontine tegmental nucleus and the laterodorsal tegmental nucleus
17-19.
Cholinergic and glutamatergic
regulation of the VTA is well established. Local application of the N-methyl-D-aspartate (NMDA) antagonist, AP5, into the rat VTA has been shown to robustly attenuate phasic DA release in the naïve rat NAc
20, 21,
SN/VTA complex
20,
learning
22.
to increase the latency to lever press for electrical self-stimulation of the
and to attenuate cue-induced cocaine seeking
21
and reward-related
Similarly, the selective genetic inactivation of NMDA receptors in mouse VTA DA
neurons attenuates both DA neuron burst firing and subsequent DA release and reward-related learning
23.
However, intra-VTA microinfusion of NMDA itself does not reliably initiate an 3 ACS Paragon Plus Environment
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increase in spontaneous DA transients in the NAc shell 20 (though it has been shown to increase the magnitude of electrically-evoked DA release in the core
24),
and when DA neurons are
recorded from slices in which afferent input has been severed, these neurons cannot be made to phasically fire in response to glutamate agonist administration alone 25, 26. Thus, it is likely that several mechanisms contribute to the generation of DA transients in the ventral striatum. Electrophysiological studies have suggested that acetylcholine plays a role in driving phasic firing in presumed VTA DA neurons
27-30.
intra-VTA self-administered carbachol
31.
This is likely critical for the reinforcing properties of
Acetylcholine receptors in the VTA have been shown
to play a key role in eliciting electrically-evoked DA release in the NAc reinforcing properties of ethanol-associated cues
32.
24
and to modulate the
Furthermore, muscarinic (but not nicotinic)
acetylcholine receptors in the VTA underlie conditioned reinforcement for a cue previously paired with food reward
33.
Collectively, these studies have begun to elucidate the role of
glutamatergic and cholinergic modulation of the VTA in conditioned reinforcement; however, far less is known about how these signals regulate the reinforcing effects of motivationally neutral sensory stimuli. Given that DA transients in the NAc are critically involved in motivation and reinforcement 34-37, that DA release in the NAc is both necessary and sufficient for cue-mediated reward-seeking behavior administration
39,
38,
and that even neutral sensory stimuli can facilitate drug self-
it is imperative to evaluate how cholinergic and glutamatergic modulation of
neuronal firing in the VTA underlies the intrinsic reinforcing effects of salient sensory stimuli that have not been previously paired with reward. In this work, we utilized fast-scan cyclic voltammetry (FSCV), which can provide selective measurements with high spatial and temporal resolution, to monitor rapidly fluctuating DA concentrations in the NAc. We monitored the effects of intra-VTA microinfusion of carbachol (non-selective cholinergic receptor agonist) by itself, or in combination with AP5 (NMDA glutamatergic receptor antagonist) or NMDA (glutamatergic receptor agonist) on the production of DA transients in the shell subregion of the NAc in freely moving animals. Additionally, we 4 ACS Paragon Plus Environment
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ACS Chemical Neuroscience
evaluated the effects of these pharmacological manipulations on a visual-stimulus-reinforced instrumental behavior task, which was designed to examine the reinforcing effects of neutral sensory stimuli. Overall, this study sheds light on the synergistic role of cholinergic and glutamatergic signaling in modulation of VTA-NAc shell dopaminergic output, and its correlation with motivation to press for a visual stimulus.
RESULTS AND DISCUSSION Cholinergic and glutamatergic mechanisms in the VTA modulate DA transients in the shell subregion of the NAc Over past two decades, FSCV has developed into a powerful tool for studying the role of DA transients in freely moving animals. These rapid DA fluctuations are thought to be the behaviorally relevant mode of DA neurotransmission, as they can become time locked to cues that predict reward availability reward
45-47.
15, 40-44,
and DA release intensifies with increasing proximity to
Studies have demonstrated DA’s role in motivation and reward-based learning, as
well as behavioral conditioning
48-52.
For instance, phasic stimulation of VTA DA neurons is
sufficient to drive behavioral conditioning to environmental context and to elicit dopamine transients in the NAc
49.
However, it remains unclear if there is a causal link between DA
signaling and the salience of an environmental cue. Thus, this study first examined the VTA mechanisms that are involved in modulation of spontaneous DA transients in the shell subregion of the NAc. Figure 1A shows a representative color plot containing 150 background-subtracted voltammograms that demonstrate DA release. Color plots allow for the discrimination of specific substances in complex brain environments, as they depict the current collected at each potential
53.
This color plot shows DA release evoked
by electrical stimulation of the VTA (asterisk), as well as a naturally occurring DA transient (pound sign). Signature cyclic voltammograms extracted from this color plot (Figure 1 C, D) serve as a means to identify DA at both time points. The current collected at the DA oxidation 5 ACS Paragon Plus Environment
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potential (0.6V – horizontal dashed line) was converted to concentration using a calibration factor, and was plotted in Figure 1B.
Figure 1. Background-subtracted voltammograms were collected every 100 msec for quantitative analysis of DA dynamics. (A) A representative color plot, containing 150 background-subtracted voltammograms that demonstrate DA release. Asterisk represents release evoked by electrical stimulation, pound represents a naturally-occurring DA transient. (B) DA concentration trace and (C, D) cyclic voltammograms extracted from the raw data presented in (A).
To examine the role that cholinergic and NMDA receptors in the VTA play in modulation of DA transients in the NAc shell, a microinfusion of saline, carbachol, a carbachol/AP5 cocktail, or a carbachol/NMDA cocktail was delivered to the posterior VTA. Immediately thereafter, the effects on DA transients recorded in the NAc (frequency and amplitude) were monitored. Figure 2A shows a representative color plot collected 2-4 minutes after microinfusion of saline (left) or carbachol (right). The DA transients are marked by white asterisks. The concentration versus 6 ACS Paragon Plus Environment
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ACS Chemical Neuroscience
time traces are shown in Figure 2B. A subsequent microinfusion of the carbachol/AP5 cocktail inhibited DA transients (Figure 2C,D left). Finally, microinfusion of a cocktail of carbachol/NMDA reinstated a robust DA signal (Figure 2C,D, right).
This unequivocally
demonstrates that the recording site still retained the ability to support DA signaling, and that the lack of signal upon administration of AP5 was not due to a broken sensor or electrode fouling.
Figure 2. Intra-VTA microinfusion of carbachol, a carbachol/AP5 cocktail, or a carbachol/NMDA cocktail modulates the frequency and amplitude of DA transients in the NAc. (A) A representative color plot collected ~3 min after microinfusion of saline (left) or carbachol (right). (B) Concentration versus time traces extracted from the data. (C) A representative color plot collected ~3 min after microinfusion of a carbachol/AP5 cocktail (left) or a carbachol/NMDA cocktail (right). (D) Concentration versus time traces extracted from the data. Asterisks represent the DA transients.
A summary of the electrochemical data is presented in Figure 3.
Figure 3A
demonstrates the effects of intra-VTA pharmacological manipulations on the frequency of DA transients recorded in the NAc, as a percent of baseline transient frequency, ([F (3, 156) = 63.90], p
