Figure 4

Macrophages express the dopamine transporter and vesicular monoamine transporter 2. Quantitative RT-PCR (A) detected mRNA for the dopamine transporter (DAT) and vesicular monoamine transporter 2 (VMAT2) in monocyte-derived macrophages (MDM) isolated from four different donors. Representative amplification curves from a single donor (A) are shown for DAT, VMAT2 (red curves) and the housekeeping gene β-actin (red curves). Amplification curves for the positive control (Total Human brain RNA, blue curves) and the negative control (cDNA negative samples, green curves) are also shown (A). Western blot analysis of isolated MDM plasma membrane (PM) and cytoplasmic (Cyto) proteins from three different donors demonstrated that DAT (B, black arrow, approximately 70 kDA) was expressed in the plasma membrane of MDM. Western blot analysis of isolated MDM total membrane (TM) and cytoplasmic (Cyto) proteins demonstrates expression of VMAT2 (4C, black arrow, approximately 48 kDA) in MDM membranes. Specificity of plasma and total membrane isolation was confirmed by enrichment of Na⁺/K⁺ ATPase and lack of α-tubulin reactivity in membrane fractions (B, C). Expression of DAT is also shown in MDM stained for DAT (4D-3, FITC) or the isotype matched negative control antibody (3D-6, FITC). Cell nuclei were stained with DAPI (4D-1, -4, blue), actin was stained with Phalloidin-Texas Red (4D-2,-5, red), and the DAT or the isotype control were stained with FITC (4D-3, 6, green). Scale bars represent 23.81 μM.