## 3 Results
### Photodynamic Opening of the BBB Using Different PSs  
In this session of experiments, we studied the PDT-opening of the BBB using different PSs, including two commercial PSs such as 5-ALA and AlPcS as well as new synthetized ones, such as ZnPc and its functionalized form GalZnPc.  
Figure 1-I illustrates molecular formulas of photosensizers used for PDT-opening of the BBB. Figure 1-II,III demonstrate confocal imaging of the BBB permeability to FITCD before and after PDT. In the control groups (no PDT, no laser or PSs), FITCD was observed only in the cerebral vessels (Figure 1-I). PDT-5-ALA/AlPcS/ZnPc/GalZnPc induced strong FITCD extravasation from the cerebral vessels into the brain tissues. The FITCD leakage was visualized by fluorescence outside the vessel walls (Figure 1-II).  
Spectrofluorimetric assay revealed laser dose-mediated increase in the BBB permeability to EBAC 1 h after PDT-5-ALA/AlPcS/ZnPc/GalZnPc. Figure 1-IV illustrates more pronounced effects of PDT-ZnPc-Gal on the EBAC leakage compared with other types of PSs (GalZnPc/AlPcS: 29.54 \(\pm\) 0.02 vs. 18.64 \(\pm\) 0.03, \(p\) \(<\) 0.001 for 2.5 J/cm\({}^{2}\); 31.80 \(\pm\) 0.03 vs. 20.37 \(\pm\) 0.08, \(p\) \(<\) 0.001 for 5 J/cm\({}^{2}\); 33.93 \(\pm\) 0.05 vs. 23.18 \(\pm\) 0.07, \(p\) \(<\) 0.001 for 10 J/cm\({}^{2}\); GalZnPc/ZnPc: 29.54 \(\pm\) 0.02 vs. 12.55 \(\pm\) 0.05, \(p\) \(<\) 0.001 for 2.5 J/cm\({}^{2}\); 31.80 \(\pm\) 0.03 vs. 14.12 \(\pm\) 0.01, \(p\) \(<\) 0.001 for 5 J/cm\({}^{2}\); 33.93 \(\pm\) 0.05 vs. 15.44 \(\pm\) 0.05, \(p\) \(<\) 0.001 for 10 J/cm\({}^{2}\); GalZnPc/5-ALA: 29.54 \(\pm\) 0.02 vs. 1.18 \(\pm\) 0.02, \(p\) \(<\) 0.001 for 2.5 J/cm\({}^{2}\); 31.80 \(\pm\) 0.03 vs. 1.09 \(\pm\) 0.02, \(p\) \(<\) 0.001 for 5 J/cm\({}^{2}\); 33.93 \(\pm\) 0.05 vs. 8.97 \(\pm\) 0.05, \(p\) \(<\) 0.001 for 10 J/cm\({}^{2}\Figure 1: Photodynamic opening of the BBB using different PSs:  
(**I**) The chemical formulas of the used PSs (A--GalZnPc; B--AlPcS; C--ZnPc; D--5-ALA) for the BBB opening; (**II**) The confocal imaging of cerebral vessels (red color) filled by FITCD (green color) in the normal state: A--the cerebral vessels are filled by FITCD (green color), B--the blood vessels marked by NG2 (pericytes marker), C--merged A and B; (**III**) The confocal imaging of extravasation of FITCD (green color, arrowed) from cerebral vessels (red color) into the brain tissues: A - the cerebral vessels are filled by FITCD (green color), B--the blood vessels marked by NG2 (pericytes marker), C--merged A and B, D and E--2-D and 3-D imaging of FITCD leakage; (**IV**) Spectrofluorimetric assay of EBAC level in the brain tissues after PDT-opening of the BBB using different PSs (5-ALA/AlPcS/ZnPc/GalZnPc): *; +\(\#\)--\(p\) < 0.001 vs. the control groups including the intact mice, mice after laser irradiation, mice treated by PSs, respectively; \(n\) = 10 for each group and \({}^{\circ}\)\(\longrightarrow\)\(<\) 0.001 between the experimental groups, at least 10 individual tissue sections taken for each animal for II and III; (**V**) The fluorescence of TJ proteins in the control group (the intact mice), 1 h and 4 h after PDT-5-ALA/AlPcS/ZnPc/GalZnPc (\(n\) = 10 for each group): **--\(p\) < 0.01, ***--\(p\) < 0.001 vs. the control group (the intact mice).