The 18-kDa mitochondrial translocator protein (TSPO) is upregulated in high-grade astrocytomas and can be imaged by PET using the selective radiotracer (11)C-(R)PK11195. We investigated (11)C-(R)PK11195 binding in human gliomas and its relationship with TSPO expression in tumor tissue and glioma-associated microglia/macrophages (GAMs) within the tumors. Twenty-two glioma patients underwent dynamic (11)C-(R)PK11195 PET scans and perfusion MR imaging acquisition. Parametric maps of (11)C-(R)PK11195 binding potential (BPND) were generated. Coregistered MR/PET images were used to guide tumor biopsy. The tumor tissue was quantitatively assessed for TSPO expression and infiltration of GAMs using immunohistochemistry and double immunofluorescence. The imaging and histopathologic parameters were compared among different histotypes and grades and correlated with each other. BPND of (11)C-(R)PK11195 in high-grade gliomas was significantly higher than in low-grade astrocytomas and low-grade oligodendrogliomas. TSPO in gliomas was expressed predominantly by neoplastic cells, and its expression correlated positively with BPND in the tumors. GAMs only partially contributed to the overall TSPO expression within the tumors, and TSPO expression in GAMs did not correlate with tumor BPND. PET with (11)C-(R)PK11195 in human gliomas predominantly reflects TSPO expression in tumor cells. It therefore has the potential to effectively stratify patients who are suitable for TSPO-targeted treatment.