Small-animal PET imaging of amyloid-beta plaques with [11C]PiB and its multi-modal validation in an APP/PS1 mouse model of Alzheimer's disease.

In vivo imaging and quantification of amyloid-ß plaque (Aß) burden in small-animal models of Alzheimer's disease (AD) is a valuable tool for translational research such as developing specific imaging markers and monitoring new therapy approaches. Methodological constraints such as image resolution of positron emission tomography (PET) and lack of suitable AD models have limited the feasibility of PET in mice. In this study, we evaluated a feasible protocol for PET imaging of Aß in mouse brain with [(11)C]PiB and specific activities commonly used in human studies. In vivo mouse brain MRI for anatomical reference was acquired with a clinical 1.5 T system. A recently characterized APP/PS1 mouse was employed to measure Aß at different disease stages in homozygous and hemizygous animals. We performed multi-modal cross-validations for the PET results with ex vivo and in vitro methodologies, including regional brain biodistribution, multi-label digital autoradiography, protein quantification with ELISA, fluorescence microscopy, semi-automated histological quantification and radioligand binding assays. Specific [(11)C]PiB uptake in individual brain regions with Aß deposition was demonstrated and validated in all animals of the study cohort including homozygous AD animals as young as nine months. Corresponding to the extent of Aß pathology, old homozygous AD animals (21 months) showed the highest uptake followed by old hemizygous (23 months) and young homozygous mice (9 months). In all AD age groups the cerebellum was shown to be suitable as an intracerebral reference region. PET results were cross-validated and consistent with all applied ex vivo and in vitro methodologies. The results confirm that the experimental setup for non-invasive [(11)C]PiB imaging of Aß in the APP/PS1 mice provides a feasible, reproducible and robust protocol for small-animal Aß imaging. It allows longitudinal imaging studies with follow-up periods of approximately one and a half years and provides a foundation for translational Alzheimer neuroimaging in transgenic mice.

Impact of the COMT Val(108/158)Met polymorphism on the mu-opioid receptor system in the human brain: mu-opioid receptor, met-enkephalin and beta-endorphin expression.

The Val(108/158)Met polymorphism of the catechol-O-methyltransferase gene (COMT) is known to interact with the function of various neuroreceptor systems in the brain. We have recently shown by post-mortem receptor autoradiography that the number of mu-opioid (MOP) receptor binding sites depends on the number of COMT Met(108/158) alleles in distinct human brain regions. We now investigated COMT Val(108/158)Met related levels of the MOP receptor protein and its endogenous ligands met-enkephalin and beta-endorphin in the human frontal cortex, thalamus and basal ganglia. Semiquantitative immunostaining and in situ hybridization were applied in a cohort of 17 human brain tissues from healthy donors. MOP receptor protein levels paralleled previous ligand binding results with a significantly higher MOP receptor expression in the mediodorsal nucleus of the thalamus of COMT Met(108/158) allele carriers. Also met-enkephalin peptide levels correlated with the genotype in this structure, with the lowest expression in COMT Met(108/158) homozygous individuals. Beta-endorphin was not detectable in the cortex, basal ganglia or thalamus, and therefore is unlikely to contribute to changes of the MOP receptor system. These results confirm the impact of the COMT Val(108/158)Met polymorphism on the MOP receptor system and may support the hypothesis of an enkephalin related turnover of MOP receptors at least in some brain structures.

Towards NR2B receptor selective imaging agents for PET-synthesis and evaluation of N-[11C]-(2-methoxy)benzyl (E)-styrene-, 2-naphthyl- and 4-trifluoromethoxyphenylamidine.

Three potent and selective 11C-labelled NR2B antagonists have been synthesized and evaluated as PET ligands. The brain uptake of the compounds in mice varied substantially and was dominated by metabolism. One compound was found to have favourable uptake and retention in the brain, as well as a binding pattern consistent with the expression of the target receptor as measured by in vitro autoradiography. However, the metabolism of the compounds tested was too rapid to allow for in vivo imaging.

Syntheses, biological evaluation, and molecular modeling of 18F-labeled 4-anilidopiperidines as mu-opioid receptor imaging agents.

The synthesis, evaluation, and molecular modeling of a series of 18F-labeled 4-anilidopiperidines with high affinities for the mu-opioid receptor (mu-OR) are reported. On the basis of the high brain uptake and selective retention in brain regions that contain a high concentration of the mu-OR, combined with a good metabolic stability, [18F]fluoro-pentyl carfentanil ([18F]4) and 2-(+/-)[18F]fluoropropyl-sufentanil ([18F]6) were selected as the lead compounds for further evaluation. The binding affinity to the human mu-OR was 0.74 and 0.13 nM for [18F]4 and [18F]6, respectively. In vitro autoradiography of [18F]4 and [18F]6 on rat brain sections produced patterns in accordance with the known distribution of mu-OR expression. Structure-activity relationships of the fluorinated compounds are discussed with respect to the interaction with an activated-state model of the mu-OR. Taken together, the in vivo and in vitro data indicate that [18F]4 and [18F]6 hold promise for studying the mu-opioid receptor in humans by means of positron emission tomography.

COMT Val108/158Met genotype affects the mu-opioid receptor system in the human brain: evidence from ligand-binding, G-protein activation and preproenkephalin mRNA expression.

Recent data from [(11)C]carfentanil ligand-PET indicate that in the human brain, the availability of mu-opioid (MOP) receptor binding sites is affected by the Val(108/158)Met polymorphism of the catechol-O-methyltransferase (COMT) gene. This prompted us to validate the impact of COMT Val(108/158)Met on MOP receptors in human post-mortem brain. [(3)H]DAMGO receptor autoradiography was performed in frontal cortex, basal ganglia, thalamus and cerebellum (8 Met/Met, 6 Met/Val, 3 Val/Val). With respect to genotype, numbers of MOP binding sites in COMT Met(108/158) homozygous and Val(108/158)Met heterozygous cases were higher than in Val(108/158) homozygous. Differences were significant in the caudate nucleus (Val/Met vs. Val/Val), nucleus accumbens (Val/Met vs. Val/Val) and the mediodorsal nucleus of the thalamus (Met/Met vs. Val/Val). In the thalamus, this was corroborated by DAMGO-stimulated [(35)S]GTPgammaS autoradiography. Moreover, stepwise multiple regression taking into account various covariables allowed to confirm the COMT genotype as the most predictive factor in this structure. As a mechanism how COMT might exert its action on MOP receptors, it has been suggested that at least in striatopallidal circuits COMT Val(108/158)Met impacts on enkephalin, which is capable of reciprocally regulating MOP receptor expression. Thus, we assessed preproenkephalin mRNA by in situ hybridization. In the striatum, mRNA levels were significantly higher in COMT Met(108/158) homozygous cases indicating that MOP binding sites and enkephalin are regulated in parallel. Moreover, the transcript was not detectable in the thalamus. Thus, mechanisms other than an enkephalin-dependent receptor turnover must be responsible for COMT-related differences in MOP binding site availability in the human brain.

18F-labeled sufentanil for PET-imaging of mu-opioid receptors.

The synthesis of an (18)F-labeled sufentanil analogue with apparent high mu-opioid receptor selectivity is reported. Intravenous injection of N-[4-(methoxymethyl)-1-[2-(2-thienyl)ethyl]-4-piperidinyl]-N-phenyl-2-(+/-)-[(18)F]fluoropropan-amide in mice resulted in high brain uptake and a regional brain activity distribution corresponding to the mu-opioid receptor expression pattern. The developed ligand is a promising tracer for extended protocols in mu-opioid receptor mapping and quantitation with positron emission tomography.

What to learn from in vivo opioidergic brain imaging?

Ligand-PET studies are attracting increasing interest in experimental and clinical research. As the most elaborated of PET techniques, ligand-PET allows the demonstration of receptor distributions, and thus, the delineation of neurochemical pathologies in the disease state. Recent developments are promising that ligand-PET will even allow to characterize dynamic and short-term changes in neurotransmission and will tremendously add to the understanding of neurophysiology on the receptor level. In pain studies, mainly the mu-opioidergic agonist [(11)C]-carfentanil and the unspecific opioid receptor antagonist [(11)C]-diprenorphine are applied. Utilizing these ligands the thalamus, prefrontal and cingulate cortex, basal ganglia and midbrain structures have been shown to possess high amounts of opioidergic receptors in vivo and it is well accepted, that the receptor density is higher in projections of the medial than those of the lateral pain system. Changes in receptor availability were observed in patients suffering from chronic pain. Rheumatoid arthritis, trigeminal neuralgia and central poststroke pain (CPSP) all lead to decreased ligand binding in pain processing regions during the painful period in comparison to pain free intervals or healthy subjects. These decreases may either be the consequence of increased endogenous release or indicate receptor internalization/down-regulation or loss of neurons carrying these receptors. Recent studies also evidenced [(11)C]-carfentanil binding changes due to acute experimental pain. One possible interpretation of these changes is that the PET-ligand might be displaced by endogenous opioidergic ligands. One major region, where this "ligand displacement" was observed, was the thalamus. These findings highlight the importance of the opioidergic system in pain processing and the power of ligand-PET to advance the understanding of pain.

SUNCT: bilateral hypothalamic activation during headache attacks and resolving of symptoms after trigeminal decompression.

Short-lasting unilateral neuralgiform headache with conjunctival injection and tearing (SUNCT) is a primary head-pain syndrome, which is often refractory to any medical treatment. Concerning the pathophysiology of SUNCT, hypothalamic involvement ipsilaterally to the pain has been suggested based on the clinical features and one functional imaging case report. Here we now report a new case with SUNCT and the concomitant cerebral activation pattern (fMRI) during the pain attacks. In addition to an activation of several brain structures known to be generally involved in pain processing, bilateral hypothalamic activation occurred during the pain attacks, arguing for a central origin of the headache. Interestingly, this patient became completely pain free after surgical decompression of the ipsilateral trigeminal nerve. We hypothesize that in this case with a central predisposition for trigeminal autonomic cephalgias, a peripheral trigger with ectopic excitation might have contributed to the clinical picture of SUNCT.

High trait anxiety and hyporeactivity to stress of the dorsomedial prefrontal cortex: a combined phMRI and Fos study in rats.

The neural basis of trait anxiety is poorly understood. In genetically selected hyperanxious (high anxiety-related behavior; HAB) rats, diazepam induces a stronger anxiolytic response than in hypoanxious (low anxiety-related behavior; LAB) rats. A screen for neuronal response differences to diazepam between HAB and LAB rats using pharmacologic fMRI (phMRI) at 7 T revealed a blunted diazepam-induced neuronal deactivation in the dorsomedial prefrontal cortex (dmPFC) of HABs. This was not due to reduced benzodiazepine (BDZ) receptor densities in this region. Instead, dmPFC tissue oxygenation at baseline was found to be significantly lower in HABs. This suggests a tonic relative hypoactivity under the highly stressful phMRI conditions, offering an explanation for the reduced responsivity to the neural depressant effect of diazepam in the sense of a floor effect. Subsequently, Fos immunoreactivity (Fos-IR) showed that ethologically relevant stressors also cause less dmPFC activation in HABs. In the context of an anxiety-inhibiting role of the dmPFC, we propose that failure to sufficiently activate this region in stressful situations may contribute to high trait anxiety.