Abstract
Rodents are popular biological models for physiological and behavioral research in neuroscience and rats are better models than mice due to their higher genome similarity to human and more accessible surgical procedures. However, rat brain is larger than mice brain and it needs powerful imaging tools to implement better penetration against the scattering of the thicker brain tissue. Three-photon fluorescence microscopy (3PFM) combined with near-infrared (NIR) excitation has great potentials for brain circuits imaging because of its abilities of anti-scattering, deep-tissue imaging, and high signal-to-noise ratio (SNR). In this work, a type of AIE luminogen with red fluorescence was synthesized and encapsulated with Pluronic F-127 to make up form nanoparticles (NPs). Bright DCDPP-2TPA NPs were employed for in vivo three-photon fluorescent laser scanning microscopy of blood vessels in rats brain under 1550[Formula: see text]nm femtosecond laser excitation. A fine three-dimensional (3D) reconstruction up to the deepness of 600[Formula: see text][Formula: see text]m was achieved and the blood flow velocity of a selected vessel was measured in vivo as well. Our 3PFM deep brain imaging method simultaneously recorded the morphology and function of the brain blood vessels in vivo in the rat model. Using this angiography combined with the arsenal of rodent’s brain disease, models can accelerate the neuroscience research and clinical diagnosis of brain disease in the future.
Highlights
We further demonstrated therst in vivo deep-tissue 3PFM imaging on rat brain by utilizing 1550 nm fs laser as the excitation, and a 600 m-deep stacked image with 5 m step, as well as three-dimensional (3D) reconstruction was acquired
After visualizing the morphology of the rat cerebral vascular system, we further measured the functional blood °ow velocity in vivo in the brain of anaesthetic rat to verify the feasibility of the Aggregation-induced emission (AIE) NPsassisted 3PFM in brain research
Each rat in the experiment group was intravenously injected 2 mL solution of DCDPP-2TPA NPs dissolved with phosphate bu®er saline (PBS) in the concentration of 1 mg mLÀ1
Summary
Multi-photon °uorescence microscopic (MPFM) imaging technologies are well known for their noninvasion and deep-penetration ability with high spatial resolution enabling scientists to visualize in vivo tissue morphology and physiology in scattering tissues.[1,2,3] Two-photon °uorescence microscopy (2PFM) and three-photon °uorescence microscopy (3PFM) combined with °uorescence labeling technology have been rapidly developed during the past few years.[4,5] Compared with 2PFM imaging, the wavelength of femtosecond (fs) laser excitation in 3PFM imaging usually is located in the second near infrared spectral region (NIR-II, 1000–1700 nm), which performs better penetration against the tissue scattering.[6,7] in the multi-photon °uorescence process, the °uorescence intensity is inversely proportional to the power of z, in which z is the distance from the focus. It is important to develop a method for in vivo rat brain imaging with high spatial resolution and large penetration depth. The mainstream diagnosis technologies of vascular system in the rat brain disease models are magnetic resonance imaging (MRI),[21] computed tomography (CT), microendoscopy,[22] photoacoustic tomographly (PAT),[23,24] etc. Compared with these modalities, 3PFM imaging has advantages of higher spatial resolution, noninvasion, quick response, and free of radiation. After visualizing the morphology of the rat cerebral vascular system, we further measured the functional blood °ow velocity in vivo in the brain of anaesthetic rat to verify the feasibility of the AIE NPsassisted 3PFM in brain research
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