Min-Liang Pan

Quantitative assessment of brown adipose tissue metabolic activity and volume using 18F-FDG PET/CT and β3-adrenergic receptor activation

BackgroundBrown adipose tissue [BAT] metabolism in vivo is vital for the development of novel strategies in combating obesity and diabetes. Currently, BAT is activated at low temperatures and measured using 2-deoxy-2-18F-fluoro-D-glucose [18F-FDG] positron-emission tomography [PET]. We report the use of β3-adrenergic receptor-mediated activation of BAT at ambient temperatures using (R, R)-5-[2-[2,3-(3-chlorphenyl)-2-hydroxyethyl-amino]propyl]-1,3-benzodioxole-2,2-dicarboxylate, disodium salt [CL316,243] (a selective β3-adrenoceptor agonist) and measured by 18F-FDG PET/computed tomography [CT].MethodsControl and CL316,243-treated (2 mg/kg) male Sprague-Dawley rats were administered with 18F-FDG for PET/CT studies and were compared to animals at cold temperatures. Receptor-blocking experiments were carried out using propranolol (5 mg/kg). Dose effects of CL316,243 were studied by injecting 0.1 to 1 mg/kg 30 min prior to 18F-FDG administration. Imaging results were confirmed by autoradiography, and histology was done to confirm BAT activation.ResultsCL316,243-activated interscapular BAT [IBAT], cervical, periaortic, and intercostal BATs were clearly visualized by PET. 18F-FDG uptake of IBAT was increased 12-fold by CL316,243 vs. 1.1-fold by cold exposure when compared to controls. 18F-FDG uptake of the CL-activated IBAT was reduced by 96.0% using intraperitoneal administration of propranolol. Average 18F-FDG uptake of IBAT increased 3.6-, 3.5-, and 7.6-fold by doses of 0.1, 0.5, and 1 mg/kg CL, respectively. Ex vivo18F-FDG autoradiography and histology of transverse sections of IBAT confirmed intense uptake in the CL-activated group and activated IBAT visualized by PET.ConclusionOur study indicated that BAT metabolic activity could be evaluated by 18F-FDG PET using CL316,243 at ambient temperature in the rodent model. This provides a feasible and reliable method to study BAT metabolism.

Adrenergic pathway activation enhances brown adipose tissue metabolism: A [18 F]FDG PET/CT study in mice

OBJECTIVE Pharmacologic approaches to study brown adipocyte activation in vivo with a potential of being translational to humans are desired. The aim of this study was to examine pre- and postsynaptic targeting of adrenergic system for enhancing brown adipose tissue (BAT) metabolism quantifiable by [(18)F]fluoro-2-deoxyglucose ([(18)F]FDG) positron emission tomography (PET)/computed tomography (CT) in mice. METHODS A β₃-adrenoreceptor selective agonist (CL 316243), an adenylyl cyclase enzyme activator (forskolin) and a potent blocker of presynaptic norepinephrine transporter (atomoxetine), were injected through the tail vein of Swiss Webster mice 30 minutes before intravenous (iv) administration of [(18)F]FDG. The mice were placed on the PET/CT bed for 30 min PET acquisition followed by 10 min CT acquisition for attenuation correction and anatomical delineation of PET images. RESULTS Activated interscapular (IBAT), cervical, periaortic and intercostal BAT were observed in 3-dimentional analysis of [(18)F]FDG PET images. CL 316243 increased the total [(18)F]FDG standard uptake value (SUV) of IBAT 5-fold greater compared to that in placebo-treated mice. It also increased the [(18)F]FDG SUV of white adipose tissue (2.4-fold), and muscle (2.7-fold), as compared to the control. There was no significant difference in heart, brain, spleen and liver uptakes between groups. Forskolin increased [(18)F]FDG SUV of IBAT 1.9-fold greater than that in placebo-treated mice. It also increased the [(18)F]FDG SUV of white adipose tissue (2.2-fold) and heart (5.4-fold) compared to control. There was no significant difference in muscle, brain, spleen, and liver uptakes between groups. Atomoxetine increased [(18)F]FDG SUV of IBAT 1.7-fold greater than that in placebo-treated mice. There were no significant differences in all other organs compared to placebo-treated mice except liver (1.6 fold increase). A positive correlation between SUV levels of IBAT and CT Hounsfield unit (HU) (R(2)=0.55, p<0.001) and between CT HU levels of IBAT and liver (R(2)=0.69, p<0.006) was observed. CONCLUSIONS The three pharmacologic approaches reported here enhanced BAT metabolism by targeting different sites in adrenergic system as measured by [(18)F]FDG PET/CT.

Striatal and Extrastriatal microPET Imaging of D2/D3 Dopamine Receptors in Rat Brain with [18F]Fallypride and [18F]Desmethoxyfallypride

In this study, we compared two different D2/3 receptor ligands, [18F]fallypride and [18F]desmethoxyfallypride ([18F]DMFP) with respect to the duration of the scan, visualization of extrastriatal receptors, and binding potentials (BPND) in the rat brain. In addition, we studied the feasibility of using these tracers following a period of awake tracer uptake, during which the animal may perform a behavioral task. Male Sprague–Dawley rats were imaged with [18F]fallypride and with [18F]DMFP in four different studies using microPET. All scans were performed under isoflurane anesthesia. The first (test) and second (retest) study were 150‐min baseline scans. No retest scans were performed with [18F]DMFP. A third study was a 60‐min awake uptake of radiotracer followed by a 90‐min scan. A fourth study was a 150‐min competition scan with haloperidol (0.2 mg/kg) administered via tail vein at 90‐min post‐[18F]fallypride injection and 60‐min post‐[18F]DMFP. For the test–retest studies, BPND was measured using both Logan noninvasive (LNI) method and the interval ratios (ITR) method. Cerebellum was used as a reference region. For the third study, the binding was measured only with the ITR method, and the results were compared to the baseline results. Studies showed that the average transient equilibrium time in the dorsal striatum (DSTR) was at 90 min for [18F]fallypride and 30 min for [18F]DMFP. The average BPND for [18F]fallypride was 14.4 in DSTR, 6.8 in ventral striatum (VSTR), 1.3 in substantia nigra/ventral tegmental area (SN/VTA), 1.4 in colliculi (COL), and 1.5 in central gray area. In the case of [18F]DMFP, the average BPND values were 2.2 in DSTR, 2.7 in VSTR, and 0.8 in SN/VTA. The haloperidol blockade showed detectable decrease in binding of both tracers in striatal regions with a faster displacement of [18F]DMFP. No significant changes in BPND of [18F]fallypride due to the initial awake state of the animal were found, whereas BPND of [18F]DMFP was significantly higher in the awake state compared to baseline. We were able to demonstrate that dynamic PET using MicroPET Inveon allows quantification of both striatal and extrastriatal [18F]fallypride binding in rats in vivo. Quantification of the striatal regions could be achieved with [18F]DMFP. Synapse 2011.

Evaluation of 18F-nifene binding to α4β2 nicotinic receptors in the rat brain using microPET imaging

MicroPET imaging studies using 18F-nifene, a new positron emission tomography (PET) radiotracer for nicotinic acetylcholinergic receptors (nAChR) α4β2 receptors in rats, have been carried out. Rats were imaged for 90 min after intravenous injection of 18F-nifene (0.8 to 1 mCi), and binding potential (BPND) was measured. 18F-Nifene binding to thalamic and extrathalamic brain regions was consistent with the α4β2 nAChR distribution in the rat brain. Using the cerebellum as a reference, the values for the thalamus varied less than 5% (BPND = 1.30, n = 3), confirming reproducibility of 18F-nifene binding. 18F-Nifene microPET imaging was also used to evaluate effects of nicotine in a group of Sprague-Dawley rats under isoflurane anesthesia. Nicotine challenge postadministration of 18F-nifene demonstrated reversibility of 18F-nifene binding in vivo. For α4β2 nAChR receptor occupancy (nAChROCC), various doses of nicotine (0, 0.02, 0.1, 0.25, and 0.50 mg/kg nicotine free base) 15 min prior to 18F-nifene were administered. Low-dose nicotine (0.02 mg) reached > 80% nAChROCC while at higher doses (0.25 mg) > 90% nAChROCC was measured. The small amount of 18F-nifene binding with reference to the cerebellum affects an accurate evaluation of nAChROCC. Efforts are underway to identify alternate reference regions for 18F-nifene microPET studies in rodents.

18F-Fallypride PET of Pancreatic Islets: In Vitro and In Vivo Rodent Studies

Islet cell loss in the pancreas results in diabetes. A noninvasive method that measures islet cell loss and also tracks the fate of transplanted islets would facilitate the development of novel therapeutics and improve the management of diabetes. We describe a novel dopamine D2/D3 receptor (D2/D3R)–based PET method to study islet cells in the rat pancreas and in islet cell transplantation. Methods: 18F-fallypride binding to isolated rat islets and pancreas was evaluated in the absence and presence of the D2/D3R inhibitor haloperidol. After intravenous 18F-fallypride (28–37 MBq) administration, normal rats and rats pretreated with haloperidol were imaged in a PET/CT scanner and subsequently studied ex vivo for 18F-fallypride localization in the pancreas. A streptozotocin-treated diabetic rat model was used to study localization of 18F-fallypride in the pancreas, in vitro and ex vivo. Rat islet cells were transplanted into the spleen and visualized using 18F-fallypride PET. Results: 18F-fallypride bound to isolated islet cells and pancreatic sections with an endocrine or exocrine selectivity of approximately 4; selectivity was reduced by haloperidol, suggesting that binding was D2/D3R-specific. Chemical destruction of islets by streptozotocin decreased 18F-fallypride binding in pancreas by greater than 50%, paralleling the decrease in insulin immunostaining. Uptake of 18F-fallypride in the pancreas was confirmed by radiochromatography and was 0.05% injected dose/cm3 as measured by PET/CT. The ratio of 18F-fallypride uptake in the pancreas to reference tissue (erector spinae muscle) was 5.5. Rat islets transplanted into the spleen were visualized in vivo by 18F-fallypride and confirmed by immunostaining. The ratio of spleen-transplanted islets to erector spinae muscle was greater than 5, compared with a ratio of 2.8 in untransplanted rats. Conclusion: These studies demonstrate the potential utility of 18F-fallypride as a PET agent for islet cells.

Dopamine D3 receptor binding of (18)F-fallypride: Evaluation using in vitro and in vivo PET imaging studies.

Identification of dopamine D3 receptors (D3R) in vivo is important to understand several brain functions related to addiction. The goal of this work was to identify D3R binding of the dopamine D2 receptor (D2R)/D3R imaging agent, 18F‐fallypride. Brain slices from male Sprague‐Dawley rats (n = 6) and New Zealand White rabbits (n = 6) were incubated with 18F‐fallypride and D3R selective agonist (R)‐7‐OH‐DPAT (98‐fold D3R selective). Rat slices were also treated with BP 897 (68‐fold D3R selective partial agonist) and NGB 2904 (56‐fold D3R selective antagonist). In vivo rat studies (n = 6) were done on Inveon PET using 18‐37 MBq 18F‐fallypride and drug‐induced displacement by (R)‐7‐OH‐DPAT, BP 897 and NGB 2904. PET/CT imaging of wild type (WT, n = 2) and D2R knock‐out (KO, n = 2) mice were carried out with 18F‐fallypride. (R)‐7‐OH‐DPAT displaced binding of 18F‐fallypride, both in vitro and in vivo. In vitro, at 10 nM (R)‐7‐OH‐DPAT, 18F‐fallypride binding in the rat ventral striatum (VST) and dorsal striatum (DST) and rabbit nucleus accumbens were reduced by ∼10–15%. At 10 μM (R)‐7‐OH‐DPAT all regions in rat and rabbit were reduced by ≥85%. In vivo reductions for DST and VST before and after (R)‐7‐OH‐DPAT were: low‐dose (0.015 mg kg−1) DST −22%, VST −29%; high‐dose (1.88 mg kg−1) DST −58%, VST −77%, suggesting D3R/D2R displacement. BP 897 and NGB 2904 competed with 18F‐fallypride in vitro, but unlike BP 897, NGB 2904 did not displace 18F‐fallypride in vivo. The D2R KO mice lacked 18F‐fallypride binding in the DST. In summary, our findings suggest that up to 20% of 18F‐fallypride may be bound to D3R sites in vivo. Synapse 69:577–591, 2015.

Targeting presynaptic norepinephrine transporter in brown adipose tissue: a novel imaging approach and potential treatment for diabetes and obesity.

Brown adipose tissue (BAT) plays a significant role in metabolism. In this study, we report the use of atomoxetine (a clinically applicable norepinephrine reuptake inhibitor) for 18F‐FDG PET imaging of BAT and its effects on heat production and blood glucose concentration. Fasted‐male Sprague‐Dawley rats were administered with intravenous 18F‐FDG. The same rats were treated with atomoxetine (0.1 mg/kg, i.v.) 30 min before 18F‐FDG administration. To confirm the β‐adrenergic effects, propranolol (β‐adrenergic inhibitor) 5 mg/kg was given intraperitoneally 30 min prior to atomoxetine administration. The effect of atomoxetine on BAT metabolism was assessed in fasted and non‐fasted rats and on BAT temperature and blood glucose in fasted rats. In 18F‐FDG PET/CT images, interscapular BAT (IBAT) and other areas of BAT were clearly visualized. When rats were fasted, atomoxetine (0.1 mg/kg) increased the 18F‐FDG uptake of IBAT by factor of 24 within 30 min. Propranolol reduced the average 18F‐FDG uptake of IBAT significantly. Autoradiography of IBAT and white adipose tissue confirmed the data obtained by PET. When rats were not fasted, atomoxetine‐induced increase of 18F‐FDG uptake in IBAT was delayed and occurred in 120 min. For comparison, direct stimulation of β3‐adrenreceptors in non‐fasted rats with CL‐316, 243 occurred within 30 min. Atomoxetine‐induced IBAT activation was associated with higher IBAT temperature and lower blood glucose. This was mediated by inhibition of norepinephrine reuptake transporters in IBAT leading to increased norepinephrine concentration in the synapse. Increased synaptic norepinephrine activates β3‐adrenreceptors resulting in BAT hypermetabolism that is visible and quantifiable by 18F‐FDG PET/CT. Synapse, 2013.

Evaluation of [18F]Nifene biodistribution and dosimetry based on whole-body PET imaging of mice.

INTRODUCTION [(18)F]Nifene is a novel radiotracer specific to the nicotinic acetylcholine α4β2 receptor class. In preparation for using this tracer in humans we have performed whole-body PET studies in mice to evaluate the in vivo biodistribution and dosimetry of [(18)F]Nifene. METHODS Seven BALB/c mice (3 males, 4 females) received IV tail injections of [(18)F]Nifene and were scanned for 2 h in an Inveon dedicated PET scanner. Each animal also received a high resolution CT scan using an Inveon CT. The CT images were used to draw volume of interest (VOI) on the following organs: brain, large intestine, small intestine, stomach, heart, kidneys, liver, lungs, pancreas, bone, spleen, testes, thymus, uterus and urinary bladder. All organ time activity curves had the decay correction reversed and were normalized to the injected activity. The area under the normalized curves was then used to compute the residence times in each organ. The absorbed doses in mouse organs were computed using the RAdiation Dose Assessment Resource (RADAR) animal models for dose assessment. The residence times in mouse organs were converted to human values using scale factors based on differences between organ and body weights. OLINDA 1.1 software was used to compute the absorbed human doses in multiple organs for both female and male phantoms. RESULTS The highest mouse residence times were found in urinary bladder, liver, bone, small intestine and kidneys. The largest doses in mice were found in urinary bladder and kidneys for both females and males. The elimination of radiotracer was primarily via kidney and urinary bladder with the urinary bladder being the limiting organ. The projected human effective doses were 1.51E-02 mSv/MBq for the adult male phantom and 1.65E-02 mSv/MBq for the adult female model phantom. CONCLUSION This study indicates that the whole-body mouse imaging can be used as a preclinical tool for initial estimation of the absorbed doses of [(18)F]Nifene in humans.

Enhancement of 18F-fluorodeoxyglucose metabolism in rat brain frontal cortex using a β3 adrenoceptor agonist.

We report the use of β3 adrenergic receptor mediated activation of rat brain frontal cortex using mirabegron (a selective β3 adrenoceptor agonist), measured by 18F‐FDG PET/CT. Another β3 agonist, CL 316,243, did not have this effect due to impermeability through the blood brain barrier (BBB), while atomoxetine, a norepinephrine transporter blocker, did increase 18F‐FDG uptake in the frontal cortex. Mirabegron exhibited a dose‐dependent increase in frontal cortex 18F‐FDG uptake. These findings suggest a possible use of selective β3 adrenoceptor agonists in reversing regional glucose hypometabolism in the brain. Synapse 69:96–98, 2015.

Imaging Pancreas in Healthy and Diabetic Rodent Model Using [18F]Fallypride Positron Emission Tomography/Computed Tomography

BACKGROUND A noninvasive method of monitoring the loss of islet cells can provide an earlier and improved diagnosis for therapeutics development of preclinical phases of diabetes. The use of [(18)F]fallypride, a dopamine D2/D3 receptor radiotracer, has been developed as a surrogate marker to evaluate loss of pancreatic islet cells in a rodent model of type 1 diabetes. MATERIALS AND METHODS Healthy Sprague-Dawley rats were administered [(18)F]fallypride and imaged for 2 h in a positron emission tomography (PET)/computed tomography (CT) scan. Diabetes was then induced in the same rats by administration of streptozotocin, and a PET/CT scan was performed 4 days after establishing diabetes. Pancreata of a separate set of rats were evaluated by insulin immunostaining for loss of islet cells by streptozotocin. RESULTS Blood glucose levels of 125 mg/dL and 550 mg/dL were established for those rats without and with diabetes, respectively. [(18)F]Fallypride uptake in the pancreas of both groups of rats was rapid, but the rats with diabetes showed a significantly lower uptake (less than 50%). The specific binding ratio was decreased by 77% in the diabetic rats. CONCLUSIONS [(18)F]Fallypride can be a useful surrogate marker for monitoring changes in pancreatic islet cells, thus providing a noninvasive method to evaluate efficacy of therapeutics.