Functional Characterization of 5-HT1B Receptor Drugs in Nonhuman Primates Using Simultaneous PET-MR.

In the present study, we used a simultaneous PET-MR experimental design to investigate the effects of functionally different compounds (agonist, partial agonist, and antagonist) on 5-HT1B receptor (5-HT1BR) occupancy and the associated hemodynamic responses. In anesthetized male nonhuman primates (n = 3), we used positron emission tomography (PET) imaging with the radioligand [11C]AZ10419369 administered as a bolus followed by constant infusion to measure changes in 5-HT1BR occupancy. Simultaneously, we measured changes in cerebral blood volume (CBV) as a proxy of drug effects on neuronal activity. The 5-HT1BR partial agonist AZ10419369 elicited a dose-dependent biphasic hemodynamic response that was related to the 5-HT1BR occupancy. The magnitude of the response was spatially overlapping with high cerebral 5-HT1BR densities. High doses of AZ10419369 exerted an extracranial tissue vasoconstriction that was comparable to the less blood-brain barrier-permeable 5-HT1BR agonist sumatriptan. By contrast, injection of the antagonist GR127935 did not elicit significant hemodynamic responses, even at a 5-HT1BR cerebral occupancy similar to the one obtained with a high dose of AZ10419369. Given the knowledge we have of the 5-HT1BR and its function and distribution in the brain, the hemodynamic response informs us about the functionality of the given drug: changes in CBV are only produced when the receptor is stimulated by the partial agonist AZ10419369 and not by the antagonist GR127935, consistent with low basal occupancy by endogenous serotonin.SIGNIFICANCE STATEMENT We here show that combined simultaneous positron emission tomography and magnetic resonance imaging uniquely enables the assessment of CNS active compounds. We conducted a series of pharmacological interventions to interrogate 5-HT1B receptor binding and function and determined blood-brain barrier passage of drugs and demonstrate target involvement. Importantly, we show how the spatial and temporal effects on brain hemodynamics provide information about pharmacologically driven downstream CNS drug effects; the brain hemodynamic response shows characteristic dose-related effects that differ depending on agonistic or antagonistic drug characteristics and on local 5-HT1B receptor density. The technique lends itself to a comprehensive in vivo investigation and understanding of drugs' effects in the brain.

News and views on in-vivo imaging of neurotransmission using PET and MRI.

Molecular neuroimaging with PET is an integrated tool in psychiatry research and drug-development for as long as this modality has been available, in particular for studying neurotransmission and endogenous neurotransmitter release. Pharmacologic, behavioral and other types of challenges are currently applied to induce changes in neurochemical levels that can be inferred through their effects on changes in receptor binding and related outcome measures. Based on the availability of tracers that are sensitive for measuring neurotransmitter release these experiments have focused on the brain's dopamine system, while recent developments have extended those studies to other targets such as the serotonin or choline system. With the introduction of hybrid, truly simultaneous PET/MRI systems, in-vivo imaging of the dynamics of neuroreceptor signal transmission in the brain using PET and functional MRI (fMRI) has become possible. fMRI has the ability to provide information about the effects of receptor function that are complementary to the PET measurement. Dynamic acquisition of both PET and fMRI signals enables not only an in-vivo real-time assessment of neurotransmitter or drug binding to receptors but also dynamic receptor adaptations and receptor-specific neurotransmission. While fMRI temporal resolution is comparatively fast in relation to PET, the timescale of observable biological processes is highly dependent on the kinetics of radiotracers and study design. Overall, the combination of the specificity of PET radiotracers to neuroreceptors, fMRI signal as a functional readout and integrated study design promises to expand our understanding of the location, propagation and connections of brain activity in health and disease.

A regularized full reference tissue model for PET neuroreceptor mapping.

The full reference tissue model (FRTM) is a PET analysis framework that includes both free and specifically bound compartments within tissues, together with rate constants defining association and dissociation from the specifically bound compartment. The simplified reference tissue model (SRTM) assumes instantaneous exchange between tissue compartments, and this "1-tissue" approximation reduces the number of parameters and enables more robust mapping of non-displaceable binding potentials. Simulations based upon FRTM have shown that SRTM exhibits biases that are spatially dependent, because biases depend upon binding potentials. In this work, we describe a regularized model (rFRTM) that employs a global estimate of the dissociation rate constant from the specifically bound compartment (k4). The model provides an internal calibration for optimizing k4 through the reference-region outflow rate k2', a model parameter that should be a global constant but varies regionally in SRTM. Estimates of k4 by rFRTM are presented for four PET radioligands. We show that SRTM introduces bias in parameter estimates by assuming an infinite value for k4, and that rFRTM ameliorates bias with an appropriate choice of k4. Theoretical considerations and simulations demonstrate that rFRTM reduces bias in non-displaceable binding potentials. A two-parameter reduction of the model (rFRTM2) provides robust mapping at a voxel-wise level. With a structure similar to SRTM, the model is easily implemented and can be applied as a PET reference region analysis that reduces parameter bias without substantially altering parameter variance.

Neurovascular coupling to D2/D3 dopamine receptor occupancy using simultaneous PET/functional MRI.

This study employed simultaneous neuroimaging with positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) to demonstrate the relationship between changes in receptor occupancy measured by PET and changes in brain activity inferred by fMRI. By administering the D2/D3 dopamine receptor antagonist [(11)C]raclopride at varying specific activities to anesthetized nonhuman primates, we mapped associations between changes in receptor occupancy and hemodynamics [cerebral blood volume (CBV)] in the domains of space, time, and dose. Mass doses of raclopride above tracer levels caused increases in CBV and reductions in binding potential that were localized to the dopamine-rich striatum. Moreover, similar temporal profiles were observed for specific binding estimates and changes in CBV. Injection of graded raclopride mass doses revealed a monotonic coupling between neurovascular responses and receptor occupancies. The distinct CBV magnitudes between putamen and caudate at matched occupancies approximately matched literature differences in basal dopamine levels, suggesting that the relative fMRI measurements reflect basal D2/D3 dopamine receptor occupancy. These results can provide a basis for models that relate dopaminergic occupancies to hemodynamic changes in the basal ganglia. Overall, these data demonstrate the utility of simultaneous PET/fMRI for investigations of neurovascular coupling that correlate neurochemistry with hemodynamic changes in vivo for any receptor system with an available PET tracer.

A 31-channel MR brain array coil compatible with positron emission tomography.

PURPOSE: Simultaneous acquisition of MR and positron emission tomography (PET) images requires the placement of the MR detection coil inside the PET detector ring where it absorbs and scatters photons. This constraint is the principal barrier to achieving optimum sensitivity on each modality. Here, we present a 31-channel PET-compatible brain array coil with reduced attenuation but improved MR sensitivity. METHODS: A series of component tests were performed to identify tradeoffs between PET and MR performance. Aspects studied include the remote positioning of preamplifiers, coax size, coil trace size/material, and plastic housing. We then maximized PET performance at minimal cost to MR sensitivity. The coil was evaluated for MR performance (signal to noise ratio [SNR], g-factor) and PET attenuation. RESULTS: The coil design showed an improvement in attenuation by 190% (average) compared with conventional 32-channel arrays, and no loss in MR SNR. Moreover, the 31-channel coil displayed an SNR improvement of 230% (cortical region of interest) compared with a PET-optimized 8-channel array with similar attenuation properties. Implementing attenuation correction of the 31-channel array successfully removed PET artifacts, which were comparable to those of the 8-channel array. CONCLUSION: The design of the 31-channel PET-compatible coil enables higher sensitivity for PET/MR imaging, paving the way for novel applications in this hybrid-imaging domain.

Dynamic functional imaging of brain glucose utilization using fPET-FDG.

Glucose is the principal source of energy for the brain and yet the dynamic response of glucose utilization to changes in brain activity is still not fully understood. Positron emission tomography (PET) allows quantitative measurement of glucose metabolism using 2-[(18)F]-fluorodeoxyglucose (FDG). However, FDG PET in its current form provides an integral (or average) of glucose consumption over tens of minutes and lacks the temporal information to capture physiological alterations associated with changes in brain activity induced by tasks or drug challenges. Traditionally, changes in glucose utilization are inferred by comparing two separate scans, which significantly limits the utility of the method. We report a novel method to track changes in FDG metabolism dynamically, with higher temporal resolution than exists to date and within a single session. Using a constant infusion of FDG, we demonstrate that our technique (termed fPET-FDG) can be used in an analysis pipeline similar to fMRI to define within-session differential metabolic responses. We use visual stimulation to demonstrate the feasibility of this method. This new method has a great potential to be used in research protocols and clinical settings since fPET-FDG imaging can be performed with most PET scanners and data acquisition and analysis are straightforward. fPET-FDG is a highly complementary technique to MRI and provides a rich new way to observe functional changes in brain metabolism.

A receptor-based model for dopamine-induced fMRI signal.

This report describes a multi-receptor physiological model of the fMRI temporal response and signal magnitude evoked by drugs that elevate synaptic dopamine in basal ganglia. The model is formulated as a summation of dopamine's effects at D1-like and D2-like receptor families, which produce functional excitation and inhibition, respectively, as measured by molecular indicators like adenylate cyclase or neuroimaging techniques like fMRI. Functional effects within the model are described in terms of relative changes in receptor occupancies scaled by receptor densities and neuro-vascular coupling constants. Using literature parameters, the model reconciles many discrepant observations and interpretations of pre-clinical data. Additionally, we present data showing that amphetamine stimulation produces fMRI inhibition at low doses and a biphasic response at higher doses in the basal ganglia of non-human primates (NHP), in agreement with model predictions based upon the respective levels of evoked dopamine. Because information about dopamine release is required to inform the fMRI model, we simultaneously acquired PET (11)C-raclopride data in several studies to evaluate the relationship between raclopride displacement and assumptions about dopamine release. At high levels of dopamine release, results suggest that refinements of the model will be required to consistently describe the PET and fMRI data. Overall, the remarkable success of the model in describing a wide range of preclinical fMRI data indicate that this approach will be useful for guiding the design and analysis of basic science and clinical investigations and for interpreting the functional consequences of dopaminergic stimulation in normal subjects and in populations with dopaminergic neuroadaptations.

A functional account of stimulation-based aerobic glycolysis and its role in interpreting BOLD signal intensity increases in neuroimaging experiments.

In aerobic glycolysis, oxygen is abundant, and yet cells metabolize glucose without using it, decreasing their ATP per glucose yield by 15-fold. During task-based stimulation, aerobic glycolysis occurs in localized brain regions, presenting a puzzle: why produce ATP inefficiently when, all else being equal, evolution should favor the efficient use of metabolic resources? The answer is that all else is not equal. We propose that a tradeoff exists between efficient ATP production and the efficiency with which ATP is spent to transmit information. Aerobic glycolysis, despite yielding little ATP per glucose, may support neuronal signaling in thin (< 0.5 µm), information-efficient axons. We call this the efficiency tradeoff hypothesis. This tradeoff has potential implications for interpretations of task-related BOLD "activation" observed in fMRI. We hypothesize that BOLD "activation" may index local increases in aerobic glycolysis, which support signaling in thin axons carrying "bottom-up" information, or "prediction error"-i.e., the BIAPEM (BOLD increases approximate prediction error metabolism) hypothesis. Finally, we explore implications of our hypotheses for human brain evolution, social behavior, and mental disorders.

A reference tissue forward model for improved PET accuracy using within-scan displacement studies.

We report a novel forward-model implementation of the full reference tissue model (fFTRM) that addresses the fast-exchange approximation employed by the simplified reference tissue model (SRTM) by incorporating a non-zero dissociation time constant from the specifically bound compartment. The forward computational approach avoided errors associated with noisy and nonorthogonal basis functions using an inverse linear model. Compared to analysis by a multilinear single-compartment reference tissue model (MRTM), fFTRM provided improved accuracy for estimation of binding potentials at early times in the scan, with no worse reproducibility across sessions. To test the model's ability to identify small focal changes in binding potential using a within-scan challenge, we employed a nonhuman primate model of focal dopamine release elicited by deep brain microstimulation remote to ventral striatum (VST) during imaging by simultaneous PET and fMRI. The new model reported an unambiguously lateralized response in VST consistent with fMRI, whereas the MRTM-derived response was not lateralized and was consistent with simulations of model bias. The proposed model enabled better accuracy in PET [11C]raclopride displacement studies and may also facilitate challenges sooner after injection, thereby recovering some sensitivity lost to radioactive decay of the PET tracer.

[11C]PBR28 radiotracer kinetics are not driven by alterations in cerebral blood flow.

The positron emission tomography (PET) radiotracer [11C]PBR28 has been increasingly used to image the translocator protein (TSPO) as a marker of neuroinflammation in a variety of brain disorders. Interrelatedly, similar clinical populations can also exhibit altered brain perfusion, as has been shown using arterial spin labelling in magnetic resonance imaging (MRI) studies. Hence, an unsolved debate has revolved around whether changes in perfusion could alter delivery, uptake, or washout of the radiotracer [11C]PBR28, and thereby influence outcome measures that affect interpretation of TSPO upregulation. In this simultaneous PET/MRI study, we demonstrate that [11C]PBR28 signal elevations in chronic low back pain patients are not accompanied, in the same regions, by increases in cerebral blood flow (CBF) compared to healthy controls, and that areas of marginal hypoperfusion are not accompanied by decreases in [11C]PBR28 signal. In non-human primates, we show that hypercapnia-induced increases in CBF during radiotracer delivery or washout do not alter [11C]PBR28 outcome measures. The combined results from two methodologically distinct experiments provide support from human data and direct experimental evidence from non-human primates that changes in CBF do not influence outcome measures reported by [11C]PBR28 PET imaging studies and corresponding interpretations of the biological meaning of TSPO upregulation.

Advances in simultaneous PET/MR for imaging neuroreceptor function.

Hybrid imaging using PET/MRI has emerged as a platform for elucidating novel neurobiology, molecular and functional changes in disease, and responses to physiological or pharmacological interventions. For the central nervous system, PET/MRI has provided insights into biochemical processes, linking selective molecular targets and distributed brain function. This review highlights several examples that leverage the strengths of simultaneous PET/MRI, which includes measuring the perturbation of multi-modal imaging signals on dynamic timescales during pharmacological challenges, physiological interventions or behavioral tasks. We discuss important considerations for the experimental design of dynamic PET/MRI studies and data analysis approaches for comparing and quantifying simultaneous PET/MRI data. The primary focus of this review is on functional PET/MRI studies of neurotransmitter and receptor systems, with an emphasis on the dopamine, opioid, serotonin and glutamate systems as molecular neuromodulators. In this context, we provide an overview of studies that employ interventions to alter the activity of neuroreceptors or the release of neurotransmitters. Overall, we emphasize how the synergistic use of simultaneous PET/MRI with appropriate study design and interventions has the potential to expand our knowledge about the molecular and functional dynamics of the living human brain. Finally, we give an outlook on the future opportunities for simultaneous PET/MRI.

Methods for Quantifying Neurotransmitter Dynamics in the Living Brain With PET Imaging.

Positron emission tomography (PET) neuroimaging in neuropsychiatry is a powerful tool for the quantification of molecular brain targets to characterize disease, assess disease subtype differences, evaluate short- and long-term effects of treatments, or even to measure neurotransmitter levels in healthy and psychiatric conditions. In this work, we present different methodological approaches (time-invariant models and models with time-varying terms) that have been used to measure dynamic changes in neurotransmitter levels induced by pharmacological or behavioral challenges in humans. The developments and potential use of hybrid PET/magnetic resonance imaging (MRI) for neurotransmission brain research will also be highlighted.

Effects of flow changes on radiotracer binding: Simultaneous measurement of neuroreceptor binding and cerebral blood flow modulation.

The potential effects of changes in blood flow on the delivery and washout of radiotracers has been an ongoing question in PET bolus injection studies. This study provides practical insight into this topic by experimentally measuring cerebral blood flow (CBF) and neuroreceptor binding using simultaneous PET/MRI. Hypercapnic challenges (7% CO2) were administered to non-human primates in order to induce controlled increases in CBF, measured with pseudo-continuous arterial spin labeling. Simultaneously, dopamine D2/D3 receptor binding of [11C]raclopride or [18F]fallypride was monitored with dynamic PET. Experiments showed that neither time activity curves nor quantification of binding through binding potentials ( BPND) were measurably affected by CBF increases, which were larger than two-fold. Simulations of experimental procedures showed that even large changes in CBF should have little effect on the time activity curves of radiotracers, given a set of realistic assumptions. The proposed method can be applied to experimentally assess the flow sensitivity of other radiotracers. Results demonstrate that CBF changes, which often occur due to behavioral tasks or pharmacological challenges, do not affect PET [11C]raclopride or [18F]fallypride binding studies and their quantification. The results from this study suggest flow effects may have limited impact on many PET neuroreceptor tracers with similar properties.

In Vivo [18F]GE-179 Brain Signal Does Not Show NMDA-Specific Modulation with Drug Challenges in Rodents and Nonhuman Primates.

As one of the major excitatory ion channels in the brain, NMDA receptors have been a leading research target for neuroscientists, physicians, medicinal chemists, and pharmaceutical companies for decades. Molecular imaging of NMDA receptors by means of positron emission tomography (PET) with [18F]GE-179 quickly progressed to clinical PET studies, but a thorough understanding of its binding specificity has been missing and has thus limited signal interpretation. Here a preclinical study with [18F]GE-179 in rodents and nonhuman primates (NHPs) is presented in an attempt to characterize [18F]GE-179 signal specificity. Rodent PET/CT was used to study drug occupancy and functional manipulation in rats by pretreating animals with NMDA targeted blocking/modulating drug doses followed by a single bolus of [18F]GE-179. Binding competition with GE-179, MK801, PCP, and ketamine, allosteric inhibition by ifenprodil, and brain activation with methamphetamine did not alter the [18F]GE-179 brain signal in rats. In addition, multimodal imaging with PET/MRI in NHPs was used to evaluate changes in radiotracer binding as a function of pharmacological challenges. Drug-induced hemodynamic changes were monitored simultaneously using functional MRI (fMRI). Comparisons of baseline and signal after drug challenge in NHPs demonstrated that the [18F]GE-179 signal cannot be manipulated in a predictable fashion in vivo. fMRI data acquired simultaneously with PET data supported this finding and provided evidence that radiotracer delivery is not altered by blood flow changes. In conclusion, the [18F]GE-179 brain signal is not readily interpretable in the context of NMDA receptor binding on the basis of the results shown in this study.

Imaging Agonist-Induced D2/D3 Receptor Desensitization and Internalization In Vivo with PET/fMRI.

This study investigated the dynamics of dopamine receptor desensitization and internalization, thereby proposing a new technique for non-invasive, in vivo measurements of receptor adaptations. The D2/D3 agonist quinpirole, which induces receptor internalization in vitro, was administered at graded doses in non-human primates while imaging with simultaneous positron emission tomography (PET) and functional magnetic resonance imaging (fMRI). A pronounced temporal divergence between receptor occupancy and fMRI signal was observed: occupancy remained elevated while fMRI responded transiently. Analogous experiments with an antagonist (prochlorperazine) and a lower-affinity agonist (ropinirole) exhibited reduced temporal dissociation between occupancy and function, consistent with a mechanism of desensitization and internalization that depends upon drug efficacy and affinity. We postulated a model that incorporates internalization into a neurovascular-coupling relationship. This model yielded in vivo desensitization/internalization rates (0.2/min for quinpirole) consistent with published in vitro measurements. Overall, these results suggest that simultaneous PET/fMRI enables characterization of dynamic neuroreceptor adaptations in vivo, and may offer a first non-invasive method for assessing receptor desensitization and internalization.

Evaluation of potential PET imaging probes for the orexin 2 receptors.

A wide range of central nervous system (CNS) disorders, particularly those related to sleep, are associated with the abnormal function of orexin (OX) receptors. Several orexin receptor antagonists have been reported in recent years, but currently there are no imaging tools to probe the density and function of orexin receptors in vivo. To date there are no published data on the pharmacokinetics (PK) and accumulation of some lead orexin receptor antagonists. Evaluation of CNS pharmacokinetics in the pursuit of positron emission tomography (PET) radiotracer development could be used to elucidate the association of orexin receptors with diseases and to facilitate the drug discovery and development. To this end, we designed and evaluated carbon-11 labeled compounds based on diazepane orexin receptor antagonists previously described. One of the synthesized compounds, [(11)C]CW4, showed high brain uptake in rats and further evaluated in non-human primate (NHP) using PET-MR imaging. PET scans performed in a baboon showed appropriate early brain uptake for consideration as a radiotracer. However, [(11)C]CW4 exhibited fast kinetics and high nonspecific binding, as determined after co-administration of [(11)C]CW4 and unlabeled CW4. These properties indicate that [(11)C]CW4 has excellent brain penetrance and could be used as a lead compound for developing new CNS-penetrant PET imaging probes of orexin receptors.