## 5 Conclusions and Outlook  
The application of PAI to biological and biomedical research has made great strides in the past twenty years. In recent years, PAI has been brought to bear on small animal brain imaging. As for PA devices, technological development is in rapid progress and the costs involved are gradually decreasing. Fiber lasers are emerging with higher pulse repetition rates and a more convenient wavelength tuning. Several groups have developed LD-PAI and LED-PAI setups that are relatively low-priced and less bulky than before. Wearable, and even wireless PAI setups, have been introduced for brain imaging, capable of observing the behaviour and activities of unanaesthetised mice, allowing for the study of the brain functions and metabolism in awake and moving subjects. This presents a new avenue for PAI-based monitoring of behaviour-related neuro and hemodynamic activities. Exogenous contrast enhancement reagents in PAI will become increasingly important for monitoring neural activity, glucose uptake, aberrant protein aggregation, malignancy, deep brain tumours, traumatic brain injury, BBB openings, epilepsy and Alzheimer's disease. Over recent years a frequent application of multi-modal imaging techniques in preclinical research has been witnessed. Examples include PAI/US for lymph nodes [63, 82], PAI/OCT (optical coherence tomography), PAI/EEG (electroencephalography) for epilepsy [85, 86, 87], PAI/PET for brain plaques [77], PAI/MRI/ FL (fluorescence) [78, 80] and PAI/MRI/computer tomography (CT) [79, 88] for tumours and glioblastomas. The skull effects arise from the acoustic speed and impedance mismatch between the skull and soft tissues, as well as from the layered structure of the skull (outer ivory-marrow-inner ivory). Speed mismatch causes phase aberration, which is the main source of artifacts in transcranial PAI. Current research shows that even a small animal skull with a thickness as thin as 0.3 mm will strongly distort PAI of the brain [83]. Three major factors that degrade the image quality of PACT are phase aberration, mode conversion and reverberation. PAM and PAMac, on the other hand, suffer mainly from signal reverberation, producing an image quality that is relatively improved and less affected by the skull. Reverberation artifacts can be easily decreased by applying low-pass filtering before the image reconstruction. However, the way in which the skull of a small animal affects the image quality may be greatly reduced by applying skull optical clearing solutions, which  
Figure 5: Simulated transcranial PAM and PACT images of a point target in water and skull cavity. (**A**) Full PACT image of the point target in water. (**B**) A close-up image of the region indicated by the dashed box in (**A**). (**C**) Full PACT image of the point target in water within the skull. (**D**) A close-up image of the region indicated by the dashed box in (**C**). (**E**) PAM image of the point source in water. (**F**) A close-up image of the region indicated by the dashed box in (**E**). (**G**) Full PAM image of the point target in water enclosed by the skull. (**H**) A close-up image of the region indicated by the dashed box in (**G**). The white dots in (**D**,**H**) represent the correct location of the point target (Reprinted with permission from Ref. [83]. 2019, Wiley-VCH). * [PERSON] et al. (2007) [PERSON]; [PERSON]; [PERSON] Photoacoustic doppler effect from flowing small light-absorbing particles. _Phys. Rev. Lett._**2007**, _99_, 184501. [CrossRef]
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