Naloxone's dose-dependent displacement of [11C]carfentanil and duration of receptor occupancy in the rat brain.

The continuous rise in opioid overdoses in the United States is predominantly driven by very potent synthetic opioids, mostly fentanyl and its derivatives (fentanyls). Although naloxone (NLX) has been shown to effectively reverse overdoses by conventional opioids, there may be a need for higher or repeated doses of NLX to revert overdoses from highly potent fentanyls. Here, we used positron emission tomography (PET) to assess NLX's dose-dependence on both its rate of displacement of [11C]carfentanil ([11C]CFN) binding and its duration of mu opioid receptor (MOR) occupancy in the male rat brain. We showed that clinically relevant doses of intravenously (IV) administered NLX (0.035 mg/kg, Human Equivalent Dose (HED) 0.4 mg; 0.17 mg/kg, HED 2 mg) rapidly displaced the specific binding of [11C]CFN in the thalamus in a dose-dependent manner. Brain MOR occupancy by IV NLX was greater than 90% at 5 min after NLX administration for both doses, but at 27.3 min after 0.035 mg/kg dose and at 85 min after 0.17 mg/kg NLX, only 50% occupancy remained. This indicates that the duration of NLX occupancy at MORs is short-lived. Overall, these results show that clinically relevant doses of IV NLX can promptly displace fentanyls at brain MORs, but repeated or higher NLX doses may be required to prevent re-narcotization following overdoses with long-acting fentanyls.

Detecting neuroinflammation in the brain following chronic alcohol exposure in rats: A comparison between in vivo and in vitro TSPO radioligand binding.

Excessive alcohol consumption is associated with neuroinflammation, which likely contributes to alcohol-related pathology. However, positron emission tomography (PET) studies using radioligands for the 18-kDa translocator protein (TSPO), which is considered a biomarker of neuroinflammation, reported decreased binding in alcohol use disorder (AUD) participants compared to controls. In contrast, autoradiographic findings in alcohol exposed rats reported increases in TSPO radioligand binding. To assess if these discrepancies reflected differences between in vitro and in vivo methodologies, we compared in vitro autoradiography (using [3 H]PBR28 and [3 H]PK11195) with in vivo PET (using [11 C]PBR28) in male, Wistar rats exposed to chronic alcohol-vapor (dependent n = 10) and in rats exposed to air-vapor (nondependent n = 10). PET scans were obtained with [11 C]PBR28, after which rats were euthanized and the brains were harvested for autoradiography with [3 H]PBR28 and [3 H]PK11195 (n = 7 dependent and n = 7 nondependent), and binding quantified in hippocampus, thalamus, and parietal cortex. Autoradiography revealed significantly higher binding in alcohol-dependent rats for both radioligands in thalamus and hippocampus (trend level for [3 H]PBR28) compared to nondependent rats, and these group differences were stronger for [3 H]PK11195 than [3 H]PBR28. In contrast, PET measures obtained in the same rats showed no group difference in [11 C]PBR28 binding. Our in vitro data are consistent with neuroinflammation associated with chronic alcohol exposure. Failure to observe similar increases in [11 C]PBR28 binding in vivo suggests the possibility that a mechanism mediated by chronic alcohol exposure interferes with [11 C]PBR28 binding to TSPO in vivo. These data question the sensitivity of PBR28 PET as a methodology to assess neuroinflammation in AUD.

Preclinical Evaluation of the First Adenosine A1 Receptor Partial Agonist Radioligand for Positron Emission Tomography Imaging.

Central adenosine A1 receptor (A1R) is implicated in pain, sleep, substance use disorders, and neurodegenerative diseases, and is an important target for pharmaceutical development. Radiotracers for A1R positron emission tomography (PET) would enable measurement of the dynamic interaction of endogenous adenosine and A1R during the sleep-awake cycle. Although several human A1R PET tracers have been developed, most are xanthine-based antagonists that failed to demonstrate competitive binding against endogenous adenosine. Herein, we explored non-nucleoside (3,5-dicyanopyridine and 5-cyanopyrimidine) templates for developing an agonist A1R PET radiotracer. We synthesized novel analogues, including 2-amino-4-(3-methoxyphenyl)-6-(2-(6-methylpyridin-2-yl)ethyl)pyridine-3,5-dicarbonitrile (MMPD, 22b), a partial A1R agonist of sub-nanomolar affinity. [11C]22b showed suitable blood-brain barrier (BBB) permeability and test-retest reproducibility. Regional brain uptake of [11C]22b was consistent with known brain A1R distribution and was blocked significantly by A1R but not A2AR ligands. [11C]22b is the first BBB-permeable A1R partial agonist PET radiotracer with the promise of detecting endogenous adenosine fluctuations.

Influence of alcoholism and cholesterol on TSPO binding in brain: PET [11C]PBR28 studies in humans and rodents.

Neuroinflammation appears to contribute to neurotoxicity observed with heavy alcohol consumption. To assess whether chronic alcohol results in neuroinflammation we used PET and [11C]PBR28, a ligand that binds to the 18-kDa translocator protein (TSPO), to compare participants with an alcohol use disorder (AUD: n = 19) with healthy controls (HC: n = 17), and alcohol-dependent (n = 9) with -nondependent rats (n = 10). Because TSPO is implicated in cholesterol's transport for steroidogenesis, we investigated whether plasma cholesterol levels influenced [11C]PBR28 binding. [11C]PBR28 binding did not differ between AUD and HC. However, when separating by TSPO genotype rs6971, we showed that medium-affinity binders AUD participants showed lower [11C]PBR28 binding than HC in regions of interest (whole brain, gray and white matter, hippocampus, and thalamus), but no group differences were observed in high-affinity binders. Cholesterol levels inversely correlated with brain [11C]PBR28 binding in combined groups, due to a correlation in AUD participants. In rodents, we observed no differences in brain [11C]PBR28 uptake between alcohol-dependent and -nondependent rats. These findings, which are consistent with two previous [11C]PBR28 PET studies, may indicate lower activation of microglia in AUD, whereas failure to observe alcohol effects in the rodent model indicate that species differences do not explain the discrepancy with prior rodent autoradiographic studies reporting increases in TSPO binding with chronic alcohol. However, reduced binding in AUD participants could also reflect competition from endogenous TSPO ligands such as cholesterol; and since the rs6971 polymorphism affects the cholesterol-binding domain of TSPO this could explain why differences were observed only in medium-affinity binders.