Continuous flushing of the bladder in rodents reduces artifacts and improves quantification in molecular imaging

In this study, we evaluated the partial volume effect (PVE) of 2-deoxy-2-[18F]fluoro-d-glucose (18F-FDG) tracer accumulation in the bladder on the positron emission tomographic (PET) image quantification in mice and rats suffering from inflammatory bowel disease. To improve the accuracy, we implemented continuous bladder flushing procedures. Female mice and rats were scanned using microPET/computed tomography (CT) at baseline and after induction of acute colitis by injecting 2,4,6-trinitrobenzene sulfonic acid (TNBS) intrarectally. During the scans, the bladder was continuously flushed in one group, whereas in the other group, no bladder flushing was performed. As a means of in vivo and ex vivo validation of the inflammation, animals also underwent colonoscopy and were sacrificed for gamma counting (subpopulation) and to score the colonic damage both micro- and macroscopically as well as biochemically. At baseline, the microPET signal in the colon of both mice and rats was significantly higher in the nonflushed group compared to the flushed group, caused by the PVE of tracer activity in the bladder. Hence, the colonoscopy and postmortem analyses showed no significant differences at baseline between the flushed and nonflushed animals. TNBS induced significant colonic inflammation, as revealed by colonoscopic and postmortem scores, which was not detected by microPET in the mice without bladder flushing, again because of spillover of bladder activity in the colonic area. MicroPET in bladder-flushed animals did reveal a significant increase in 18F-FDG uptake. Correlations between microPET and colonoscopy, macroscopy, microscopy, and myeloperoxidase yielded higher Spearman rho values in mice with continuously flushed bladders during imaging. Comparable, although somewhat less pronounced, results were shown in the rat. Continuous bladder flushing reduced image artifacts and is mandatory for accurate image quantification in the pelvic region for both mice and rats. We designed and validated experimental protocols to facilitate such.Steven Deleye, Marthe Heylen, Annemie Deiteren, Joris De Man, Sigrid Stroobants, Benedicte De Winter, and Steven Staelen

Absence of cardiovascular manifestations in a haploinsufficient Tgfbr1 mouse model

Loeys-Dietz syndrome (LDS) is an autosomal dominant arterial aneurysm disease belonging to the spectrum of transforming growth factor β (TGFβ)-associated vasculopathies. In its most typical form it is characterized by the presence of hypertelorism, bifid uvula/cleft palate and aortic aneurysm and/or arterial tortuosity. LDS is caused by heterozygous loss of function mutations in the genes encoding TGFβ receptor 1 and 2 (TGFBR1 and -2), which lead to a paradoxical increase in TGFβ signaling. To address this apparent paradox and to gain more insight into the pathophysiology of aneurysmal disease, we characterized a new Tgfbr1 mouse model carrying a p.Y378*nonsense mutation. Study of the natural history in this model showed that homozygous mutant mice die during embryonic development due to defective vascularization. Heterozygous mutant mice aged 6 and 12 months were morphologically and (immuno)histochemically indistinguishable from wild-type mice. We show that the mutant allele is degraded by nonsense mediated mRNA decay, expected to result in haploinsufficiency of the mutant allele. Since this haploinsufficiency model does not result in cardiovascular malformations, it does not allow further study of the process of aneurysm formation. In addition to providing a comprehensive method for cardiovascular phenotyping in mice, the results of this study confirm that haploinsuffciency is not the underlying genetic mechanism in human LDS

A multimodal post processing framework for activation studies of the rat brain

A stimulus-on versus stimulus-off imaging study is often used to evaluate the brain's response to a presented visual, electrical or chemical trigger. Clinical software and human templates already exist but given the recent advent of ultrahigh resolution μSPECT and μPET, a larger need rises for a post processing platform to perform these subtraction molecular imaging studies also in small animals. We have designed such a multimodal framework to perform μSPECT activation studies in rats thereby making use of μCT and MRI for anatomical land marking. METHODS: Our software solution is a combination of Amide, MRIcroN and a custom made Matlab implementation. We have studied the performance of different deep brain stimulations for which 6 rats were implanted with a multi-polar stimulation electrode in the right hippocampus. Each animal underwent a 99mTc-HMPAO μSPECT with the Milabs U-SPECT-II and a μCT scan with the GMI X-O CT before and after stimulation. Two line markers in oblique positions filled with low activity of 125I are used to register the μSPECT and μCT images. Afterwards the animals were sacrificed, their electrode was removed and a MRI scan was performed using the wrist coil of a Siemens Trio 3T. RESULTS: A semi-automated five step procedure delivers the activation map: (i) first the stimulus-on μCT and the MRI are registered to the stimulus-off μCT followed by (ii) the fusion of the off/on μSPECT scans with their off/on μCT counterparts. From the MRI, (iii) the rat brain is extracted, which is used as a mask for the calculation. Afterwards, (iv) both off/on μSPECT scans are normalized and subtracted within this MRI brain mask. Finally, (v) the Z-score, representing the activation map, is achieved by dividing the result with the standard deviation of the masked stimulus-off μSPECT. CONCLUSION: This semi-automated approach allows the experimental neuroscientist to draw conclusions on the location, spatial extent and intensity of the small animal brain’s response to the stimulus.status: publishe

Radiosynthesis and in vivo evaluation of [11C]-labelled pyrrole-2-carboxamide derivates as novel radioligands for PET imaging of monoamine oxidase A

Introduction: Since MAO-A is an enzyme involved in the metabolism of neurotransmitters, fluctuations in MAO-A functionality are associated with psychiatric and neurological disorders as well as with tobacco addiction and behaviour. This study reports the radiolabelling of two [11C]-labelled pyrrole-2-carboxamide derivates, RS 2315 and RS 2360, along with the characterization of their in vivo properties. Methods: The radiolabelling of [11C]-RS 2315 and [11C]-RS 2360 was accomplished by alkylation of their amide precursors with [11C]CH3I. Biodistribution, blocking and metabolite studies of both tracers were performed in NMRI mice. Finally, a PET study in Sprague-Dawley rats was performed for [11C]-RS 2360. Results: Both tracers were obtained in a radiochemical yield of approximately 30% with radiochemical purity of >98%. Biodistribution studies showed high brain uptake followed by rapid brain clearance for both radiotracers. In the brain, [11C]-RS 2360 was more stable than [11C]-RS 2315. Blocking studies in mice could not demonstrate specificity of [11C]-RS 2315 towards MAO-A or MAO-B. The blocking and imaging study with [11C]-RS 2360 on the other hand indicated specific binding in MAO-A at the earliest time points. Conclusions: [11C]-RS 2315 displayed a high nonspecific binding and is therefore not suitable for visualization of MAO-A in vivo. [11C]-RS 2360 on the other hand has potential for mapping MAO-A since specific binding is demonstrated. © 2010

Hippocampal deep brain stimulation induces decreased rCBF in the hippocampal formation of the rat

Deep brain stimulation (DBS) is a promising experimental approach to treat various neurological disorders. However, the optimal stimulation paradigm and the precise mechanism of action of DBS are unknown. Neuro-imaging by means of Single Photon Emission Computed Tomography (SPECT) is a non-invasive manner of evaluating regional cerebral blood flow (rCBF) changes, which are assumed to reflect changes in neural activity. In this study, rCBF changes induced by hippocampal DBS are evaluated by subtraction analysis of stimulation on/off using small animal microSPECT of the rat brain. Rats (n=13) were implanted with a multi-contact DBS electrode in the right hippocampus and injected with 10 mCi of HMPAO-Tc99(m) during application of various hippocampal DBS paradigms and amplitudes and during sham stimulation. Subtraction analysis revealed that hippocampal DBS caused a significant decrease in relative rCBF, both in the ipsi- (the side of the implanted electrode) and contralateral hippocampus. Hypoperfusion spread contralaterally with increasing stimulation amplitude. A clear distinction in spatial extent and intensity of hypoperfusion was observed between stimulation paradigms: bipolar Poisson Distributed Stimulation induced significant hypoperfusion ipsi- and contralaterally (p<0.01), while during other stimulation paradigms, rCBF-changes were less prominent. In conclusion, small animal microSPECT allows us to draw conclusions on the location, spatial extent and intensity of the hypoperfusion observed in the ipsi- and contralateral hippocampus, induced by hippocampal DBS. Our study demonstrates an innovative approach to visualize the effects of DBS and can be a useful tool in evaluating the effect of various stimulation paradigms and target areas for DBS

The Effects of Physiological and Methodological Determinants on 18

Introduction: In this study, the influence of physiological determinants on 18F-fluoro- d -glucose ( 18 F-FDG) brain uptake was evaluated in a mouse model of Alzheimer disease. Materials and Methods: TASTPM (Tg) and age-matched C57BL/6 J (WT) mice were fasted for 10 hours, while another group was fasted for 20 hours to evaluate the effect of fasting duration. The effect of repeatedly scanning was evaluated by scanning Tg and WT mice at days 1, 4, and 7. Brain 18 F-FDG uptake was evaluated in the thalamus being the most indicative region. Finally, the cerebellum was tested as a reference region for the relative standard uptake value (rSUV). Results: When correcting the brain uptake for glucose, the effect of different fasting durations was attenuated and the anticipated hypometabolism in Tg mice was demonstrated. Also, with repeated scanning, the brain uptake values within a group and the hypometabolism of the Tg mice only remained stable over time when glucose correction was applied. Finally, hypometabolism was also observed in the cerebellum, yielding artificially higher rSUV values for Tg mice. Conclusion: Corrections for blood glucose levels have to be applied when semiquantifying 18 F-FDG brain uptake in mouse models for AD. Potential reference regions for normalization should be thoroughly investigated to ensure that they are not pathologically affected also by afferent connections