Quantification of Cerebral Vascular Architecture using Two-photon Microscopy in a Mouse Model of HIV-induced Neuroinflammation

Abstract

Human Immunodeficiency Virus 1 (HIV-1) infection frequently results in HIV-1 Associated Neurocognitive Disorders (HAND), and is characterized by a chronic neuroinflammatory state within the central nervous system (CNS), thought to be driven principally by virally-mediated activation of microglia and brain resident macrophages. HIV-1 infection is also accompanied by changes in cerebrovascular blood flow (CBF), raising the possibility that HIV-associated chronic neuroinflammation may lead to changes in CBF and/or in cerebral vascular architecture. To address this question, we have used a mouse model for HIV-induced neuroinflammation, and we have tested whether long-term exposure to this inflammatory environment may damage brain vasculature and result in rarefaction of capillary networks. In this paper we describe a method to quantify changes in cortical capillary density in a mouse model of neuroinflammatory disease (HIV-1 Tat transgenic mice). This generalizable approach employs in vivo two-photon imaging of cortical capillaries through a thin-skull cortical window, as well as ex vivo two-photon imaging of cortical capillaries in mouse brain sections. These procedures produce images and z-stack files of capillary networks, respectively, which can be then subjected to quantitative analysis in order to assess changes in cerebral vascular architecture. Video LinkThe video component of this article can be found at

A quantitative brain map of experimental cerebral malaria pathology

“…Thus, although the presentation of pRBC accumulation in the cerebrovasculature may be different in ECM and HCM, our results provide a rational potential explanation for the comparable alterations in blood flow observed in vivo during murine and human malaria-induced cerebral pathology [ 54 – 59 ]. Whether the differential natures of pRBC-mediated vascular occlusion during ECM and HCM depend upon established differences in murine and human capillary diameter (average 3um vs 6.4um) [ 60 – 62 ], or upon the degree to which murine and human cerebral capillaries can mechanically dilate, requires further investigation.…”

Microglia at center stage: a comprehensive review about the versatile and unique residential macrophages of the central nervous system

“…First experimental therapeutic approaches have been suggested [ 208 ], but more research is required. Substantial research of this topic has used live animal imaging [ 209 , 210 ], often the retina model, as the blood vessels of the eye are easily monitored. However, more in vivo research about the influence of microglia on the BBB would be needed and new models ought to be developed.…”

Intermittent compression increases flow through a gel: a mechanical model perhaps relevant to flow of cerebrospinal fluid

“…Q = 0.86 × 10 −7 µL/min = 1.4 fL/s = 1.4 × 10 −18 m 3 /s (2) Taking the external diameter of a brain capillary as 4 µm [1, 58, 61] and the thickness of the basement lamina as 0.1 µm [62-64], the cross-section of the basement lamina of a capillary is …”

“…The total length of capillaries per unit volume of brain (in mouse, rat and cat) is about 1000 mm/mm 3 [1,[58][59][60] and the mean length of individual capillaries is about 100 µm [2]. Hence, there are 10 4 capillaries per mm 3 or 10 7 per cm 3 and the paracapillary flow of CSF along each capillary is Q = 0.86 x 10 -7 µL/min = 1.4 fL/s = 1.4 x 10 -18 m 3 /s (2) Taking the external diameter of a brain capillary as 4 µm [1,58,61] and the thickness of the basement lamina as 0.1 µm [62][63][64], the cross-section of the basement lamina of a capillary is A =  x (diameter) x (thickness) = 4 x 0.1 µm 2 = 1.26 µm 2…”

“…The total length of capillaries per unit volume of brain (in mouse, rat and cat) is about 1000 mm/mm 3 [1,[62][63][64] and the mean length of individual capillaries is about 100 µm [2]. Hence, there are 10 4 capillaries per mm 3 or 10 7 per cm 3 and the paracapillary flow of CSF along each capillary is Q = 0.86 x 10 -7 µL/min = 1.4 fL/s = 1.4 x 10 -18 m 3 /s (2) Taking the external diameter of a brain capillary as 4 µm [1,62,65] and the thickness of the basement lamina as 0.1 µm [66][67][68], the cross-section of the basement lamina of a capillary is A =  x (diameter) x (thickness) = 4 x 0.1 µm 2 = 1.26 µm 2 (3) Speed of flow along basement lamina = Q/A = 1.4 x 10 -18 m 3 s -1 /(1.26 x 10 -12 m 2 ) = 1.11 µm/s (4) The data in Fig. 2C show that for frequencies in the range of the mouse heart beat ( 7 -12 Hz; [53] and a pressure gradient of 40 cm H2O over 0.4 cm, dx/dt in the microburette  40 µm/s.…”