Detecting and quantifying c-fos in vivo by PET imaging of DREADD-expressing neurons in mice brains after sleep deprivation

• Nachweis und Quantifizierung von c-fos in vivo durch PET-Bildgebung von DREADD-exprimierenden Neuronen im Gehirn von Mäusen nach Schlafentzug

He, Xuan; Kampa, Björn M. (Thesis advisor); Müller, Frank (Thesis advisor); Elmenhorst, David (Thesis advisor)

Aachen : RWTH Aachen University (2021)
Dissertation / PhD Thesis

Dissertation, RWTH Aachen University, 2021

Abstract

Sleep is a critical but fragile behaviour which includes many complex and unresolved questions such as the specific cellular and molecular mechanisms of sleep homeostasis. In this thesis, in order to locate and identify the cellular populations particularly affected by sleep deprivation in mice, we developed a in vivo novel system combining Designer Receptor Exclusively Activated by Designer Drugs (DREADDs) technique with positron emission tomography (PET) method in double transgenic hM3Dqfos mice (cfos-controlled DREADDs). In this system, c-fos proteins activated by sleep deprivation was determined as the marker of neural activity indirectly via quantification of DREADDs with [11C]clozapine and [11C]CNO PET imaging in vivo. After quantitative evaluation of the [11C]clozapine and [11C]CNO in vivo binding results with binding potential (BPND), total distribution volume (VT), as well as standard uptake values (SUVs), we indicated the brain regions which were significantly affected by sleep deprivation based on both atlas and statistical parametric mapping (SPM) analysis technique. However, due to the test-retest reliability evaluation and potential in vivo binding in cerebellum for BPND and SUVAUC of CB data, the values from VT and SUVAUC of IDIF were recommended for performing quantification analysis of [11C]clozapine and [11C]CNO PET imaging with better reproducibility and reliability. Moreover, in order to improve the brain uptake of [11C]CNO, we explored to apply tariquidar before intravenous injection of the radioligand. Nevertheless, according to the in vivo quantification analysis, [11C]clozapine did not present higher affinity to DREADDs compared with [11C]CNO regarding to VT and SUVAUC of IDIF. In addition, the in vitro immunohistochemistry experiments were conducted to confirm the construct of double transgenic (hM3Dqfos) mouse model as well as the quantification evaluation of [11C]clozapine and [11C]CNO PET imaging.On the other hand, in order to guarantee a good condition for mice and reduce the stress as much as possible during the longitudinal study with DREADD and PET, an image-derived input function (IDIF) instead of the arterial input function method was applied for PET quantification based on the results from Chapter 3. In that methodology chapter (Chapter 3), we established, evaluated and selected the best non-invasive IDIF method based on the PET imaging for the A1 adenosine receptors (A1ARs) with [18F]CPFPX. Here, three different IDIF approaches were selected for evaluation. Subsequently, a VOI over the heart became the best choice to extract the IDIF after comparing with another two different VOI placements regarding image-derived curves, area under curve (AUC) ratios, and kinetic modelling results. In Chapter 3, correction of partial volume effect and metabolism were investigated as well with [18F]CPFPX. Finally, the IDIF method over heart (cube, 8 mm) were utilized in evaluation of c-fos expression by imaging DREADDs with [11C]clozapine and [11C]CNO as PET radioligands.