The small GTP-binding protein Rab8 is known to play an essential role in intracellular transport and cilia formation. We have previously demonstrated that Rab8a is required for localising apical markers in various organisms. Rab8a has a closely related isoform, Rab8b. To determine whether Rab8b can compensate for Rab8a, we generated Rab8b-knockout mice. Although the Rab8b-knockout mice did not display an overt phenotype, Rab8a and Rab8b double-knockout mice exhibited mislocalisation of apical markers and died earlier than Rab8a-knockout mice. The apical markers accumulated in three intracellular patterns in the double-knockout mice. However, the localisation of basolateral and/or dendritic markers of the double-knockout mice seemed normal. The morphology and the length of various primary and/or motile cilia, and the frequency of ciliated cells appeared to be identical in control and double-knockout mice. However, an additional knockdown of Rab10 in double-knockout cells greatly reduced the percentage of ciliated cells. Our results highlight the compensatory effect of Rab8a and Rab8b in apical transport, and the complexity of the apical transport process. In addition, neither Rab8a nor Rab8b are required for basolateral and/or dendritic transport. However, simultaneous loss of Rab8a and Rab8b has little effect on ciliogenesis, whereas additional loss of Rab10 greatly affects ciliogenesis.

The norepinephrine transporter (NET) substrates [ 123 I]meta-iodobenzylguanidine (MIBG) and [ 11 C] meta-hydroxyephedrine (HED) are used as markers of cardiac sympathetic neurons and adrenergic tumors (pheochromocytoma, neuroblastoma). However, their rapid NET transport rates limit their ability to provide accurate measurements of cardiac nerve density. [ 11 C]Phenethylguanidine ([ 11 C] 1a) and 12 analogs ([ 11 C]1b-m) were synthesized and evaluated as radiotracers with improved kinetics for quantifying cardiac nerve density. In isolated rat hearts, neuronal uptake rates of [ 11 C] 1a-m ranged from 0.24 to 1.96 mL/min/g wet, and six compounds had extremely long neuronal retention times (clearance T 1/2 > 20 hr) due to efficient vesicular storage. PET studies in nonhuman primates with [ 11 C]1e, N-[ 11 C]guanyl-meta-octopamine, which has a slow NET transport rate, showed improved myocardial kinetics compared to HED. Compound [ 11 C]1c, [ 11 C]parahydroxyphenethylguandine, which has a rapid NET transport rate, avidly accumulated into rat pheochromocytoma xenograft tumors in mice. These encouraging findings demonstrate that radiolabeled phenethylguanidines deserve further investigation as radiotracers of cardiac sympathetic innervation and adrenergic tumors.

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4-[18F]fluoro-m-hydroxyphenethylguanidine ([18F]4F-MHPG, [18F]1) is a new cardiac sympathetic nerve radiotracer with kinetic properties favorable for quantifying regional nerve density with PET and tracer kinetic analysis. An automated synthesis of [18F]1 was developed in which the intermediate 4-[18F]fluoro-m-tyramine ([18F]16) was prepared using a diaryliodonium salt precursor for nucleophilic aromatic [18F]fluorination. In PET imaging studies in rhesus macaque monkeys, [18F]1 demonstrated high quality cardiac images with low uptake in lungs and liver. Compartmental modeling of [18F]1 kinetics provided ‘net uptake rate’ constants Ki (mL/min/g wet) and Patlak graphical analysis of [18F]1 kinetics provided Patlak slopes Kp (mL/min/g). In pharmacological blocking studies with the norepinephrine transporter inhibitor desipramine (DMI), each of these quantitative measures declined in a dose-dependent manner with increasing DMI doses. These initial results strongly suggest that [18F]1 can provide quantitative measures of regional cardiac sympathetic nerve density in human hearts using PET.

The small GTP-binding protein Rab8 is known to play an essential role in intracellular transport and cilia formation. We have previously demonstrated that Rab8a is required for localising apical markers in various organisms. Rab8a has a closely related isoform, Rab8b. To determine whether Rab8b can compensate for Rab8a, we generated Rab8b-knockout mice. Although the Rab8b-knockout mice did not display an overt phenotype, Rab8a and Rab8b double-knockout mice exhibited mislocalisation of apical markers and died earlier than Rab8a-knockout mice. The apical markers accumulated in three intracellular patterns in the double-knockout mice. However, the localisation of basolateral and/or dendritic markers of the double-knockout mice seemed normal. The morphology and the length of various primary and/or motile cilia, and the frequency of ciliated cells appeared to be identical in control and double-knockout mice. However, an additional knockdown of Rab10 in double-knockout cells greatly reduced the percentage of ciliated cells. Our results highlight the compensatory effect of Rab8a and Rab8b in apical transport, and the complexity of the apical transport process. In addition, neither Rab8a nor Rab8b are required for basolateral and/or dendritic transport. However, simultaneous loss of Rab8a and Rab8b has little effect on ciliogenesis, whereas additional loss of Rab10 greatly affects ciliogenesis.