Publications using the RZ5P

Hunger increases the motivation for calorie consumption, often at the expense of low taste appeal. However, the neural mechanisms integrating calorie-sensing with increased motivation for calorie consumption remain unknown. Agouti-related peptide neurons in the arcuate nucleus of the hypothalamus sense hunger, and the ingestion of caloric solutions promote dopamine release in the absence of sweet taste perception. Therefore, we hypothesized that metabolic-sensing of hunger by AgRP neurons would be essential to promote dopamine release in the nucleus accumbens in response to caloric, but not non-caloric solutions. Moreover, we examined whether metabolic sensing in AgRP neurons affected taste preference to bitter solutions under conditions of energy need. Here we show that impaired metabolic sensing in AgRP neurons attenuated nucleus accumbens dopamine release in response to sucrose, but not saccharin, consumption. Further, metabolic sensing in AgRP neurons was essential to distinguish nucleus accumbens dopamine response to sucrose consumption when compared with saccharin. Under conditions of hunger, metabolic sensing in AgRP neurons increased the preference of sucrose solutions laced with the bitter tastant, quinine, to ensure calorie consumption whereas mice with impaired metabolic sensing in AgRP neurons maintained a strong aversion to sucrose/quinine solutions despite ongoing hunger. In conclusion, we demonstrate normal metabolic sensing in AgRP neurons drives the preference for calorie consumption, primarily when needed, by engaging dopamine release in nucleus accumbens.

The rewarding taste of food is critical for motivating animals to eat, but whether taste has a parallel function in promoting meal termination is not well understood. Here we show that hunger-promoting AgRP neurons are rapidly inhibited during each bout of ingestion by a signal linked to the taste of food. Blocking these transient dips in activity via closed-loop optogenetic stimulation increases food intake by selectively delaying the onset of satiety. We show that upstream leptin receptor-expressing neurons in the dorsomedial hypothalamus (DMH LepR ) are tuned to respond to sweet or fatty tastes and exhibit time-locked activation during feeding that is the mirror image of downstream AgRP cells. These findings reveal an unexpected role for taste in the negative feedback control of ingestion. They also reveal a mechanism by which AgRP neurons, which are the primary cells that drive hunger, are able to influence the moment-by-moment dynamics of food consumption.

Objective: The sensory detection of food and food cues suppresses Agouti related peptide (AgRP) neuronal activity prior to consumption with greatest suppression occurring in response to highly caloric food or interoceptive energy need. However, the interoceptive mechanisms priming an appropriate AgRP neural response to external sensory information of food availability remain unexplored. Since hunger increases plasma ghrelin, we hypothesized that ghrelin receptor (GHSR) signalling on AgRP neurons is a key interoceptive mechanism integrating energy need with external sensory cues predicting caloric availability. Methods: We used in vivo photometry to measure the effects of ghrelin administration or fasting on AgRP neural activity with GCaMP6s and dopamine release in the nucleus accumbens with GRAB-DA in mice lacking ghrelin receptors in AgRP neurons. Results: The deletion of GHSR on AgRP neurons prevented ghrelin-induced food intake, motivation and AgRP activity. The presentation of food (peanut butter pellet) or a wooden dowel suppressed AgRP activity in fasted WT but not mice lacking GHSRs in AgRP neurons. Similarly, peanut butter and a wooden dowel increased dopamine release in the nucleus accumbens after ip ghrelin injection in WT but not mice lacking GHSRs in AgRP neurons. No difference in dopamine release was observed in fasted mice. Finally, ip ghrelin administration did not directly increase dopamine neural activity in the ventral tegmental area. Conclusions: Our results suggest that AgRP GHSRs integrate an interoceptive state of energy need with external sensory information to produce an optimal change in AgRP neural activity. Thus, ghrelin signalling on AgRP neurons is more than just a feedback signal to increase AgRP activity during hunger.

The termination of a meal is controlled by dedicated neural circuits in the caudal brainstem. A key challenge is to understand how these circuits transform the sensory signals generated during feeding into dynamic control of behaviour. The caudal nucleus of the solitary tract (cNTS) is the first site in the brain where many meal-related signals are sensed and integrated 1–4 , but how the cNTS processes ingestive feedback during behaviour is unknown. Here we describe how prolactin-releasing hormone (PRLH) and GCG neurons, two principal cNTS cell types that promote non-aversive satiety, are regulated during ingestion. PRLH neurons showed sustained activation by visceral feedback when nutrients were infused into the stomach, but these sustained responses were substantially reduced during oral consumption. Instead, PRLH neurons shifted to a phasic activity pattern that was time-locked to ingestion and linked to the taste of food. Optogenetic manipulations revealed that PRLH neurons control the duration of seconds-timescale feeding bursts, revealing a mechanism by which orosensory signals feed back to restrain the pace of ingestion. By contrast, GCG neurons were activated by mechanical feedback from the gut, tracked the amount of food consumed and promoted satiety that lasted for tens of minutes. These findings reveal that sequential negative feedback signals from the mouth and gut engage distinct circuits in the caudal brainstem, which in turn control elements of feeding behaviour operating on short and long timescales.

Adaptive behavior in a dynamic environment often requires rapid revaluation of stimuli that deviates from well-learned associations. The divergence between stable value-encoding and appropriate behavioral output remains a critical test to theories of dopamine’s function in learning, motivation, and motor control. Yet how dopamine neurons are involved in the revaluation of cues when the world changes to alter our behavior remains unclear. Here we make use of pharmacology, in vivo electrophysiology, fiber photometry, and optogenetics to resolve the contributions of the mesolimbic dopamine system to the dynamic reorganization of reward-seeking. Male and female rats were trained to discriminate when a conditioned stimulus would be followed by sucrose reward by exploiting the prior, non-overlapping presentation of a separate discrete cue - an occasion setter. Only when the occasion setter’s presentation preceded the conditioned stimulus did the conditioned stimulus predict sucrose delivery. As a result, in this task we were able to dissociate the average value of the conditioned stimulus from its immediate expected value on a trial-to-trial basis. Both the activity of ventral tegmental area dopamine neurons and dopamine signaling in the nucleus accumbens were essential for rats to successfully update behavioral responding in response to the occasion setter. Moreover, dopamine release in the nucleus accumbens following the conditioned stimulus only occurred when the occasion setter indicated it would predict reward. Downstream of dopamine release, we found that single neurons in the nucleus accumbens dynamically tracked the value of the conditioned stimulus. Together these results reveal a novel mechanism within the mesolimbic dopamine system for the rapid revaluation of motivation.

Tyrosine hydroxylase (TH)-containing neurons of the dopamine (DA) cell group A13 are well positioned to impact known DA-related functions as their descending projections innervate target regions that regulate vigilance, sensory integration, and motor execution. Despite this connectivity, little is known regarding the functionality of A13-DA circuits. Using TH-specific loss-of-function methodology and techniques to monitor population activity in transgenic rats in vivo, we investigated the contribution of A13-DA neurons in reward and movement-related actions. Our work demonstrates a role for A13-DA neurons in grasping and handling of objects but not reward. A13-DA neurons responded strongly when animals grab and manipulate food items, whereas their inactivation or degeneration prevented animals from successfully doing so-a deficit partially attributed to a reduction in grip strength. By contrast, there was no relation between A13-DA activity and food-seeking behavior when animals were tested on a reward-based task that did not include a reaching/grasping response. Motivation for food was unaffected, as goal-directed behavior for food items was in general intact following A13 neuronal inactivation/degeneration. An anatomical investigation confirmed that A13-DA neurons project to the superior colliculus (SC) and also demonstrated a novel A13-DA projection to the reticular formation (RF). These results establish a functional role for A13-DA neurons in prehensile actions that are uncoupled from the motivational factors that contribute to the initiation of forelimb movements and help position A13-DA circuits into the functional framework regarding centrally located DA populations and their ability to coordinate movement.

In the classical model of the basal ganglia, direct pathway striatal projection neurons (dSPNs) send projections to the substantia nigra (SNr) and entopeduncular nucleus to regulate motor function. Recent studies have re-established that dSPNs also possess axon collaterals within the globus pallidus (GPe) (bridging collaterals), yet the significance of these collaterals for behavior is unknown. Here we use in vivo optical and chemogenetic tools combined with deep learning approaches in mice to dissect the roles of dSPN GPe collaterals in motor function. We find that dSPNs projecting to the SNr send synchronous motor-related information to the GPe via axon collaterals. Inhibition of native activity in dSPN GPe terminals impairs motor activity and function via regulation of Npas1 neurons. We propose a model by which dSPN GPe axon collaterals (striatopallidal Go pathway) act in concert with the canonical terminals in the SNr to support motor control by inhibiting Npas1 neurons.

Rapid-eye-movement (REM) sleep is accompanied by intense cortical activity, underlying its wake-like electroencephalogram (EEG). The neural activity inducing REM sleep is thought to originate from subcortical circuits in brainstem and hypothalamus. However, whether cortical neurons can also trigger REM sleep has remained unknown. Here, we show in mice that the medial prefrontal cortex (mPFC) strongly promotes REM sleep. Bidirectional optogenetic manipulations demonstrate that excitatory mPFC neurons promote REM sleep through their projections to the lateral hypothalamus (LH) and regulate phasic events, reflected in accelerated EEG theta oscillations and increased eye-movement density during REM sleep. Calcium imaging reveals that the majority of LH-projecting mPFC neurons are maximally activated during REM sleep and a subpopulation is recruited during phasic theta accelerations. Our results delineate a cortico-hypothalamic circuit for the top-down control of REM sleep and identify a critical role of the mPFC in regulating phasic events during REM sleep.

The circadian rhythm pacemaker, the suprachiasmatic nucleus (SCN), mediates light entrainment via vasoactive intestinal peptide (VIP) neurons (SCNVIP). Yet, how these neurons uniquely respond and connect to intrinsically photosensitive retinal ganglion cells (ipRGCs) expressing melanopsin (Opn4) has not been determined functionally in freely behaving animals. To address this, we first used monosynaptic tracing from SCNVIP neurons in mice and identified two SCNVIP subpopulations. Second, we recorded calcium changes in response to ambient light, at both bulk and single-cell levels, and found two unique activity patterns in response to high- and low-intensity blue light. The activity patterns of both subpopulations could be manipulated by application of an Opn4 antagonist. These results suggest that the two SCNVIP subpopulations connect to two types of Opn4-expressing ipRGCs, likely M1 and M2, but only one is responsive to red light. These findings have important implications for our basic understanding of non-image-forming circadian light processing.

The N -acyl phosphatidylethanolamine-specific phospholipase D (NAPE-PLD) catalyzes the production of N -acylethanolamines (NAEs), a family of endogenous bioactive lipids, which are involved in various biological processes ranging from neuronal functions to energy homeostasis and feeding behaviors. Reward-dependent behaviors depend on dopamine (DA) transmission between the ventral tegmental area (VTA) and the nucleus accumbens (NAc), which conveys reward-values and scales reinforced behaviors. However, whether and how NAPE-PLD may contribute to the regulation of feeding and reward-dependent behaviors has not yet been investigated. This biological question is of paramount importance since NAEs are altered in obesity and metabolic disorders. Here, we show that transcriptomic meta-analysis highlights a potential role for NAPE-PLD within the VTA®NAc circuit. Using brain-specific invalidation approaches, we report that the integrity of NAPE-PLD is required for the proper homeostasis of NAEs within the midbrain VTA and it affects food-reward behaviors. Moreover, region-specific knock-down of NAPE-PLD in the VTA enhanced food-reward seeking and reinforced behaviors, which were associated with increased in vivo DA release dynamics in response to both food and non-food-related rewards together with heightened tropism towards food consumption. Furthermore, midbrain knock-down of NAPE-PLD, which increased energy expenditure and adapted nutrient partitioning, elicited a relative protection against high-fat diet-mediated body fat gain and obesity-associated metabolic features. In conclusion, these findings reveal a new key role of VTA NAPE-PLD in shaping DA-dependent events, feeding behaviors and energy homeostasis, thus providing new insights on the regulation of body metabolism.

Aims Major depressive disorder is a severe psychiatric disorder that afflicts ~17% of the world population. Neuroimaging investigations of depressed patients have consistently reported the dysfunction of the basolateral amygdala in the pathophysiology of depression. However, how the BLA and related circuits are implicated in the pathogenesis of depression is poorly understood. Methods Here, we combined fiber photometry, immediate early gene expression (c‐fos), optogenetics, chemogenetics, behavioral analysis, and viral tracing techniques to provide multiple lines of evidence of how the BLA neurons mediate depressive‐like behavior. Results We demonstrated that the aversive stimuli elevated the neuronal activity of the excitatory BLA neurons (BLA CAMKII neurons). Optogenetic activation of CAMKII neurons facilitates the induction of depressive‐like behavior while inhibition of these neurons alleviates the depressive‐like behavior. Next, we found that the chemogenetic inhibition of GABAergic neurons in the BLA (BLA GABA ) increased the firing frequency of CAMKII neurons and mediates the depressive‐like phenotypes. Finally, through fiber photometry recording and chemogenetic manipulation, we proved that the activation of BLA GABA neurons inhibits BLA CAMKII neuronal activity and alleviates depressive‐like behavior in the mice. Conclusion Thus, through evaluating BLA GABA and BLA CAMKII neurons by distinct interaction, the BLA regulates depressive‐like behavior.

Rapid-eye-movement sleep (REMs) is characterized by activated electroencephalogram (EEG) and muscle atonia, accompanied by vivid dreams. REMs is homeostatically regulated, ensuring that any loss of REMs is compensated by a subsequent increase in its amount. However, the neural mechanisms underlying the homeostatic control of REMs are largely unknown. Here, we show that GABAergic neurons in the preoptic area of the hypothalamus projecting to the tuberomammillary nucleus (POA GAD2 →TMN neurons) are crucial for the homeostatic regulation of REMs. POA GAD2 →TMN neurons are most active during REMs, and inhibiting them specifically decreases REMs. REMs restriction leads to an increased number and amplitude of calcium transients in POA GAD2 →TMN neurons, reflecting the accumulation of REMs pressure. Inhibiting POA GAD2 →TMN neurons during REMs restriction blocked the subsequent rebound of REMs. Our findings reveal a hypothalamic circuit whose activity mirrors the buildup of homeostatic REMs pressure during restriction and that is required for the ensuing rebound in REMs.

Degeneration of dopamine neurons in the substantia nigra and their striatal axon terminals causes cardinal motor symptoms of Parkinson’s disease (PD). In idiopathic cases, high levels of mitochondrial DNA (mtDNA) mutations associated with mitochondrial dysfunction are a central feature of these vulnerable neurons. Here we present a mouse model expressing the K320E-variant of the mitochondrial helicase Twinkle in dopamine neurons, leading to accelerated mtDNA ageing. K320E-Twinkle DaN mice showed normal motor function at 20 months of age, although already ∼70% of nigral dopamine neurons had perished. The remaining neuron population still preserved ∼75% of axon terminals in the dorsal striatum, which enabled normal dopamine release. Transcriptome analysis and viral tracing confirmed compensatory axonal sprouting of surviving nigral dopamine neurons. We conclude that a small population of substantia nigra neurons can adapt to mtDNA mutations and maintain motor control in mice, holding chances for new treatment strategies in PD patients.

To investigate whether glucoregulatory neurons in the hypothalamus can sense and respond to physiological variation in the blood glucose (BG) level, we combined continuous arterial glucose monitoring with continuous measures of the activity of a specific subset of neurons located in the hypothalamic ventromedial nucleus that express pituitary adenylate cyclase activating peptide (VMNPACAP neurons) obtained using fiber photometry. Data were collected in conscious, free-living mice during a 1-h baseline monitoring period and a subsequent 2-h intervention period during which the BG level was raised either by consuming a chow or a high-sucrose meal or by intraperitoneal glucose injection. Cross-correlation analysis revealed that, following a 60- to 90-s delay, interventions that raise the BG level reliably associate with reduced VMNPACAP neuron activity (P < 0.01). In addition, a strong positive correlation between BG and spontaneous VMNPACAP neuron activity was observed under basal conditions but with a much longer (∼25 min) temporal offset, consistent with published evidence that VMNPACAP neuron activation raises the BG level. Together, these findings are suggestive of a closed-loop system whereby VMNPACAP neuron activation increases the BG level; detection of a rising BG level, in turn, feeds back to inhibit these neurons. To our knowledge, these findings constitute the first evidence of a role in glucose homeostasis for glucoregulatory neurocircuits that, like pancreatic β-cells, sense and respond to physiological variation in glycemia. Article Highlights By combining continuous arterial glucose monitoring with fiber photometry, studies investigated whether neurons in the murine ventromedial nucleus that express pituitary adenylate cyclase activating peptide (VMNPACAP neurons) detect and respond to changes in glycemia in vivo. VMNPACAP neuron activity rapidly decreases (within <2 min) when the blood glucose level is raised by either food consumption or glucose administration. Spontaneous VMNPACAP neuron activity also correlates positively with glycemia, but with a longer temporal offset, consistent with reports that hyperglycemia is induced by experimental activation of these neurons. Like pancreatic β-cells, neurons in the hypothalamic ventromedial nucleus appear to sense and respond to physiological variation in glycemia.

We introduce a new concept for reverse engineering silent neuronal networks using a supervised learning algorithm combined with stimulation. We quantify the performance of the algorithm and the precision of deriving synaptic weights in inhibitory and excitatory subpopulations. We then show that stimulation enables deciphering connectivity of heterogeneous circuits fed with real electrode array recordings, which could extend in the future to deciphering connectivity in broad biological and artificial neural networks.

Midbrain and striatal dopamine signals have been extremely well characterized over the past several decades, yet novel dopamine signals and functions in reward learning and motivation continue to emerge. A similar characterization of real-time sub-second dopamine signals in areas outside of the striatum has been limited. Recent advances in fluorescent sensor technology and fiber photometry permit the measurement of dopamine binding correlates, which can divulge basic functions of dopamine signaling in non-striatal dopamine terminal regions, like the dorsal bed nucleus of the stria terminalis (dBNST). Here, we record GRAB DA signals in the dBNST during a Pavlovian lever autoshaping task. We observe greater Pavlovian cue-evoked dBNST GRAB DA signals in sign-tracking (ST) compared to goal-tracking/intermediate (GT/INT) rats and the magnitude of cue-evoked dBNST GRAB DA signals decreases immediately following reinforcer-specific satiety. When we deliver unexpected rewards or omit expected rewards, we find that dBNST dopamine signals encode bidirectional reward prediction errors in GT/INT rats, but only positive prediction errors in ST rats. Since sign- and goal-tracking approach strategies are associated with distinct drug relapse vulnerabilities, we examined the effects of experimenter-administered fentanyl on dBNST dopamine associative encoding. Systemic fentanyl injections do not disrupt cue discrimination but generally potentiate dBNST dopamine signals. These results reveal multiple dBNST dopamine correlates of learning and motivation that depend on the Pavlovian approach strategy employed.

Reward-predictive cues acquire motivating and reinforcing properties that contribute to the escalation and relapse of drug use in addiction. The ventral pallidum (VP) and ventral tegmental area (VTA) are two key nodes in brain reward circuitry implicated in addiction and necessary for the performance of cue-driven behavior. Evidence suggests that VP neurons projecting to the VTA (VP→VTA) promote cue-induced reinstatement of drug-seeking, but the mechanisms by which these neurons do so are undefined. In addition, the role of these neurons in the pursuit of non-drug reward is not known. In the current study, we used in vivo fiber photometry and optogenetics to record from and manipulate VP→VTA in rats performing a discriminative stimulus task (DS task) with sucrose reward to determine the fundamental role these neurons play in invigoration and reinforcement by reward and associated discriminative cues. We find that VP→VTA neurons are selectively active during reward consumption, that optogenetic stimulation of these neurons paired with reward consumption biases choice, and that VP→VTA optogenetic stimulation is reinforcing. Critically, we found no significant encoding of cue-elicited reward-seeking vigor and acute optogenetic stimulation of these neurons paired with cue onset did not enhance the probability or vigor of reward-seeking. Our results suggest that VP→VTA neurons are active during the consumption of natural reward and that this activity reinforces seeking behavior.

The medial prefrontal cortex (mPFC) regulates cognitive flexibility and emotional behavior. Neurons that release serotonin project to the mPFC, and serotonergic drugs influence emotion and cognition. Yet, the specific roles of endogenous serotonin release in the mPFC on neurophysiology and behavior are unknown. We show that axonal serotonin release in the mPFC directly inhibits the major mPFC output neurons. In serotonergic neurons projecting from the dorsal raphe to the mPFC, we find endogenous activity signatures pre-reward retrieval and at reward retrieval during a cognitive flexibility task. In vivo optogenetic activation of this pathway during pre-reward retrieval selectively improved extradimensional rule shift performance while inhibition impaired it, demonstrating sufficiency and necessity for mPFC serotonin release in cognitive flexibility. Locomotor activity and anxiety-like behavior were not affected by either optogenetic manipulation. Collectively, our data reveal a powerful and specific modulatory role of endogenous serotonin release from dorsal raphe-to-mPFC projecting neurons in cognitive flexibility.

SignificanceGenetically encoded optical probes to image calcium levels in neurons in vivo are used widely as a real-time measure of neuronal activity in the brain. Mesoscale calcium imaging through a cranial window provides a method of studying the interaction of circuit activity between cortical areas but lacks access to subcortical regions.AimWe have developed an optical and surgical preparation that preserves wide-field imaging of the cortical surface while also permitting access to specific subcortical networks.ApproachThis was achieved using an optical fiber implanted in the striatum, along with a bilateral widefield cranial window, enabling simultaneous mesoscale cortical imaging and subcortical fiber photometry recording of calcium signals in a transgenic animal expressing GCaMP. Subcortical signals were collected from the dorsal regions of the striatum. We combined this approach with multiple sensory-motor tasks, including specific auditory and visual stimulation, and video monitoring of animal movements and pupillometry during head-fixed behaviors.ResultsWe found high correlations between cortical and striatal activity in response to sensory stimulation or movement. Furthermore, spontaneous activity recordings revealed that specific motifs of cortical activity are correlated with presynaptic activity recorded in the striatum, enabling us to select for corticostriatal activity motifs.ConclusionWe believe that this method can be utilized to reveal not only global patterns but also cell-specific connectivity that provides insight into corticobasal ganglia circuit organization.

We examined the effect of neonatal deafening on frequency-specific pathways for processing of interaural time differences (ITDs) in cochlear-implant stimuli. Animal studies have demonstrated differences in neural ITD sensitivity in the inferior colliculus (IC) depending on the intracochlear location of intracochlear stimulating electrodes. We used neonatally deafened (ND) rats of both sexes and recorded the responses of single neurons in the IC to electrical stimuli with ITDs delivered to the apical or basal cochlea and compared them with acutely deafened (AD) rats of both sexes with normal hearing (NH) during development. We found that neonatal deafness significantly impacted the ITD sensitivity and the ITD tuning patterns restricted to apically driven IC neurons. In ND rats, the ITD sensitivity of apically driven neurons is reduced to values similar to basally driven neurons. The prevalence of ITD-sensitive apical neurons with a peak-shaped ITD tuning curve, which may reflect predominant input from the medial superior olivary (MSO) complex, in ND rats was diminished compared with that in AD rats (67%, AD vs 40%, ND). Conversely, monotonic-type responses rarely occurred in AD rats (14%) but were approximately equally as prevalent as peak-type tuning curves in ND rats (42%). Nevertheless, in ND rats, the ITD at the maximum slope of the ITD tuning curve was still more concentrated within the physiological ITD range in apically driven than in basally driven neurons. These results indicate that the development of high ITD sensitivity processed by low-frequency pathways depends on normal auditory experience and associated biases in ITD tuning strategies.