The chemogenetic technology referred to as designer receptors exclusively activated by designer drugs (DREADDs) offers reversible means to control neuronal activity for investigating its functional ...correlation with behavioral action. Deschloroclozapine (DCZ), a recently developed highly potent and selective DREADD actuator, displays a capacity to expand the utility of DREADDs for chronic manipulation without side effects in nonhuman primates, which has not yet been validated. Here we investigated the pharmacokinetics and behavioral effects of orally administered DCZ in female and male macaque monkeys. Pharmacokinetic analysis and PET occupancy examination demonstrated that oral administration of DCZ yielded slower and prolonged kinetics, and that its bioavailability was 10%-20% of that in the case of systemic injection. Oral DCZ (300-1000 μg/kg) induced significant working memory impairments for at least 4 h in monkeys with hM4Di expressed in the dorsolateral prefrontal cortex (Brodmann's area 46). Repeated daily oral doses of DCZ consistently caused similar impairments over two weeks without discernible desensitization. Our results indicate that orally delivered DCZ affords a less invasive strategy for chronic but reversible chemogenetic manipulation of neuronal activity in nonhuman primates, and this has potential for clinical application.
The use of designer receptors exclusively activated by designer drugs (DREADDs) for chronic manipulation of neuronal activity for days to weeks may be feasible for investigating brain functions and behavior on a long time-scale, and thereby for developing therapeutics for brain disorders, such as epilepsy. Here we performed pharmacokinetics and
occupancy study of orally administered deschloroclozapine to determine a dose range suitable for DREADDs studies. In monkeys expressing hM4Di in the prefrontal cortex, single and repeated daily doses significantly induced working-memory impairments for hours and over two weeks, respectively, without discernible desensitization. These results indicate that orally delivered deschloroclozapine produces long-term stable chemogenetic effects, and holds great promise for the translational use of DREADDs technology.
The chemogenetic technology designer receptors exclusively activated by designer drugs (DREADDs) afford remotely reversible control of cellular signaling, neuronal activity and behavior. Although the ...combination of muscarinic-based DREADDs with clozapine-N-oxide (CNO) has been widely used, sluggish kinetics, metabolic liabilities and potential off-target effects of CNO represent areas for improvement. Here, we provide a new high-affinity and selective agonist deschloroclozapine (DCZ) for muscarinic-based DREADDs. Positron emission tomography revealed that DCZ selectively bound to and occupied DREADDs in both mice and monkeys. Systemic delivery of low doses of DCZ (1 or 3 μg per kg) enhanced neuronal activity via hM3Dq within minutes in mice and monkeys. Intramuscular injections of DCZ (100 μg per kg) reversibly induced spatial working memory deficits in monkeys expressing hM4Di in the prefrontal cortex. DCZ represents a potent, selective, metabolically stable and fast-acting DREADD agonist with utility in both mice and nonhuman primates for a variety of applications.
The term 'temporal discounting' describes both choice preferences and motivation for delayed rewards. Here we show that neuronal activity in the dorsal part of the primate caudate head (dCDh) signals ...the temporally discounted value needed to compute the motivation for delayed rewards. Macaque monkeys performed an instrumental task, in which visual cues indicated the forthcoming size and delay duration before reward. Single dCDh neurons represented the temporally discounted value without reflecting changes in the animal's physiological state. Bilateral pharmacological or chemogenetic inactivation of dCDh markedly distorted the normal task performance based on the integration of reward size and delay, but did not affect the task performance for different reward sizes without delay. These results suggest that dCDh is involved in encoding the integrated multi-dimensional information critical for motivation.
It has been widely accepted that dopamine (DA) plays a major role in motivation, yet the specific contribution of DA signaling at D
1
-like receptor (D
1
R) and D
2
-like receptor (D
2
R) to ...cost–benefit trade-off remains unclear. Here, by combining pharmacological manipulation of DA receptors (DARs) and positron emission tomography (PET) imaging, we assessed the relationship between the degree of D
1
R/D
2
R blockade and changes in benefit- and cost-based motivation for goal-directed behavior of macaque monkeys. We found that the degree of blockade of either D
1
R or D
2
R was associated with a reduction of the positive impact of reward amount and increasing delay discounting. Workload discounting was selectively increased by D
2
R antagonism. In addition, blocking both D
1
R and D
2
R had a synergistic effect on delay discounting but an antagonist effect on workload discounting. These results provide fundamental insight into the distinct mechanisms of DA action in the regulation of the benefit- and cost-based motivation, which have important implications for motivational alterations in both neurological and psychiatric disorders.
Positron emission tomography (PET) allows biomolecular tracking but PET monitoring of brain networks has been hampered by a lack of suitable reporters. Here, we take advantage of bacterial ...dihydrofolate reductase, ecDHFR, and its unique antagonist, TMP, to facilitate in vivo imaging in the brain. Peripheral administration of radiofluorinated and fluorescent TMP analogs enabled PET and intravital microscopy, respectively, of neuronal ecDHFR expression in mice. This technique can be used to the visualize neuronal circuit activity elicited by chemogenetic manipulation in the mouse hippocampus. Notably, ecDHFR‐PET allows mapping of neuronal projections in non‐human primate brains, demonstrating the applicability of ecDHFR‐based tracking technologies for network monitoring. Finally, we demonstrate the utility of TMP analogs for PET studies of turnover and self‐assembly of proteins tagged with ecDHFR mutants. These results establish opportunities for a broad spectrum of previously unattainable PET analyses of mammalian brain circuits at the molecular level.
SYNOPSIS
ecDHFR‐based reporter system can be utilized for bimodal fluorescence and Positron emission tomography (PET) imaging of expression and dynamics of its fused protein of interest in living animal brains, offering broad‐spectrum analyses of a mammalian deep brain circuit at molecular levels.
We established a genetically encoded ecDHFR‐based reporter system applicable for bimodal optical and PET imaging in living animal brains.
The reporter gene expression driven by an activity‐dependent promoter illuminates neuronal ensemble activities elicited by chemogenetic manipulation in the mouse hippocampal circuit.
ecDHFR/TMP systems enable visualization of neuronal tracts in deep brain regions of non‐human primates.
The utility of TMP analogs for PET monitoring of aggregation and turnover of proteins tagged with mutant forms of ecDHFR.
Application of bacterial dihydrofolate reductase ecDHFR and its unique antagonist TMP achieves a broad spectrum of previously unattainable in vivo PET analyses of mammalian brain circuits at the molecular level.
Serotonin (5-HT) deficiency is a core biological pathology underlying depression and other psychiatric disorders whose key symptoms include decreased motivation. However, the exact role of 5-HT in ...motivation remains controversial and elusive. Here, we pharmacologically manipulated the 5-HT system in macaque monkeys and quantified the effects on motivation for goal-directed actions in terms of incentives and costs. Reversible inhibition of 5-HT synthesis increased errors and reaction times on goal-directed tasks, indicating reduced motivation. Analysis found incentive-dependent and cost-dependent components of this reduction. To identify the receptor subtypes that mediate cost and incentive, we systemically administered antagonists specific to 4 major 5-HT receptor subtypes: 5-HT1A, 5-HT1B, 5-HT2A, and 5-HT4. Positron emission tomography (PET) visualized the unique distribution of each subtype in limbic brain regions and determined the systemic dosage for antagonists that would achieve approximately 30% occupancy. Only blockade of 5-HT1A decreased motivation through changes in both expected cost and incentive; sensitivity to future workload and time delay to reward increased (cost) and reward value decreased (incentive). Blocking the 5-HT1B receptor also reduced motivation through decreased incentive, although it did not affect expected cost. These results suggest that 5-HT deficiency disrupts 2 processes, the subjective valuation of costs and rewards, via 5-HT1A and 5-HT1B receptors, thus leading to reduced motivation.
Epilepsy is a disorder in which abnormal neuronal hyperexcitation causes several types of seizures. Because pharmacological and surgical treatments occasionally interfere with normal brain function, ...a more focused and on-demand approach is desirable. Here we examined the efficacy of a chemogenetic tool-designer receptors exclusively activated by designer drugs (DREADDs)-for treating focal seizure in a nonhuman primate model. Acute infusion of the GABA
receptor antagonist bicuculline into the forelimb region of unilateral primary motor cortex caused paroxysmal discharges with twitching and stiffening of the contralateral arm, followed by recurrent cortical discharges with hemi- and whole-body clonic seizures in two male macaque monkeys. Expression of an inhibitory DREADD (hM4Di) throughout the seizure focus, and subsequent on-demand administration of a DREADD-selective agonist, rapidly suppressed the wide-spread seizures. These results demonstrate the efficacy of DREADDs for attenuating cortical seizure in a nonhuman primate model.
The orbitofrontal cortex (OFC) and its major downstream target within the basal ganglia-the rostromedial caudate nucleus (rmCD)-are involved in reward-value processing and goal-directed behavior. ...However, a causal contribution of the pathway linking these two structures to goal-directed behavior has not been established. Using the chemogenetic technology of designer receptors exclusively activated by designer drugs with a crossed inactivation design, we functionally and reversibly disrupted interactions between the OFC and rmCD in two male macaque monkeys. We injected an adeno-associated virus vector expressing an inhibitory designer receptor, hM4Di, into the OFC and contralateral rmCD, the expression of which was visualized
by positron emission tomography and confirmed by postmortem immunohistochemistry. Functional disconnection of the OFC and rmCD resulted in a significant and reproducible loss of sensitivity to the cued reward value for goal-directed action. This decreased sensitivity was most prominent when monkeys had accumulated a certain amount of reward. These results provide causal evidence that the interaction between the OFC and the rmCD is needed for motivational control of action on the basis of the relative reward value and internal drive. This finding extends the current understanding of the physiological basis of psychiatric disorders in which goal-directed behavior is affected, such as obsessive-compulsive disorder.
In daily life, we routinely adjust the speed and accuracy of our actions on the basis of the value of expected reward. Abnormalities in these kinds of motivational adjustments might be related to behaviors seen in psychiatric disorders such as obsessive-compulsive disorder. In the current study, we show that the connection from the orbitofrontal cortex to the rostromedial caudate nucleus is essential for motivational control of action in monkeys. This finding expands our knowledge about how the primate brain controls motivation and behavior and provides a particular insight into disorders like obsessive-compulsive disorder in which altered connectivity between the orbitofrontal cortex and the striatum has been implicated.
Dump trucks are widely used in the construction industry. When sand is discharged from a dump truck bed, the friction with the bed causes sand to remain on the bed. Therefore, it is desirable to ...develop a technology to adhere low-friction fluororesin to the steel plates of a truck bed. This study develops a technique for directly joining dissimilar materials, namely steel and polytetrafluoroethylene (PTFE), using one-sided seam welding and investigates the effects of heat input and pressure conditions on joining quality. The joining conditions that affect joining quality are examined. The results show that tensile shear strength tends to increase with increasing heat input. Furthermore, it is found that the amount of PTFE deformation and the fracture morphology during tensile testing change with roller pressurization. For a one-sided seam welding machine, the anchor effect occurs between the electrodes with roller pressurization but does not occur without roller pressurization. It is also found that the type of anchor effect (dense or dispersed) depends on whether roller pressure is applied. Furthermore, it is suggested that the amount of deformation of PTFE during tensile testing is different caused by the different form of occurrence of the anchor effect. It was assumed that the fracture morphology of PTFE changed as the amount of deformation increased during the tensile test, approaching the elongation limit of PTFE. Furthermore, when the rollers were cooled to increase the cooling function of the rollers, the number of anchors was found to be lower than when the rollers were not cooled. This result suggested that the thermal contraction of the PTFE during cooling causes the PTFE in the concave area to peel off. From these results, it is assumed that the reason for the dispersed anchor effect is due to the influence of heat removal by the roller pressurization.
To interrogate particular neuronal pathways in nonhuman primates under natural and stress-free conditions, we applied designer receptors exclusively activated by designer drugs (DREADDs) technology ...to common marmosets. We injected adeno-associated virus vectors expressing the excitatory DREADD hM3Dq into the unilateral substantia nigra (SN) in four marmosets. Using multi-tracer positron emission tomography imaging, we detected DREADD expression in vivo, which was confirmed in nigrostriatal dopamine neurons by immunohistochemistry, as well as by assessed activation of the SN following agonist administration. The marmosets rotated in a contralateral direction relative to the activated side 30–90 min after consuming food containing the highly potent DREADD agonist deschloroclozapine (DCZ) but not on the following days without DCZ. These results indicate that non-invasive and reversible DREADD manipulation will extend the utility of marmosets as a primate model for linking neuronal activity and natural behavior in various contexts.
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•DREADDs were applied to activate unilateral nigrostriatal dopamine neurons•DREADD expression/function and marmoset behavior were monitored non-invasively•Orally delivered DREADD actuator reversibly induced contralateral rotation behavior•Non-invasive chemogenetic manipulation will broaden the utility of marmosets
Behavioral neuroscience; Molecular neuroscience; Cellular neuroscience
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