Intrathecal antibody distribution in the rat brain: surface diffusion, perivascular transport and osmotic enhancement of delivery

Abstract

The precise mechanisms governing the central distribution of macromolecules from the cerebrospinal fluid (CSF) to the brain and spinal cord remain poorly understood, despite their importance for physiological processes such as antibody trafficking for central immune surveillance, as well as several ongoing intrathecal clinical trials. In the present study, we clarify how IgG and smaller single-domain antibodies (sdAb) distribute throughout the whole brain in a size-dependent manner after intrathecal infusion in rats using ex vivo fluorescence and in vivo three-dimensional magnetic resonance imaging. Antibody distribution was characterized by diffusion at the brain surface and widespread distribution to deep brain regions along the perivascular spaces of all vessel types, with sdAb accessing a four- to seven-fold greater brain area than IgG. Perivascular transport involved blood vessels of all caliber and putative smooth muscle and astroglial basement membrane compartments. Perivascular access to smooth muscle basement membrane compartments also exhibited size-dependence. Electron microscopy was used to show stomata on leptomeningeal coverings of blood vessels in the subarachnoid space as potential access points allowing substances in the CSF to enter the perivascular space. Osmolyte co-infusion significantly enhanced perivascular access of the larger antibody from the CSF, with intrathecal 0.75 m mannitol increasing the number of perivascular profiles per slice area accessed by IgG by ∼50%. The results of the present study reveal potential distribution mechanisms for endogenous IgG, which is one of the most abundant proteins in the CSF, as well as provide new insights with respect to understanding and improving the drug delivery of macromolecules to the central nervous system via the intrathecal route.

The role of brain barriers in fluid movement in the CNS: is there a ‘glymphatic’ system?

“…Potential routes of entry from the CSF into the PVS include specialized pores, termed stomata, recently demonstrated on the adventitial lining cells of leptomeningeal vessels in the SAS of the rat by scanning electron microscopy [137] (Fig. 1b), confirming earlier decades-old identification of such structures in cats [199]; similar pores may also exist on the pia [30,146], providing an additional route into the PVS via the subpial space (discussed in [137]). It has become increasingly clear that substances within the CSF may potentially access and distribute along the PVS to varying extents all throughout the cerebrovascular tree, e.g., large full length antibodies (immunoglobulin G) have been shown to access the PVS of arterioles (Fig.…”

“…1c), capillaries (Fig. 1d), and venules following intrathecal infusion in rats [137]. The demonstration that CSF-infused tracers can distribute perivascularly even along microvessels comprising the classical NVU has now led to a number of interesting questions regarding (1) how such a perivascular distribution might accomplish CSF-ISF exchange, (2) the roles that astrocytes may play in regulating CSF-ISF exchange, and (3) the transport processes (e.g., diffusion, dispersion, or convection) governing fluid and tracer movement within the PVS and the surrounding brain ECS (Fig.…”

“…2). The PVS is unquestionably a site of great importance due to its involvement in disease processes affecting the NVU (e.g., cerebral amyloid angiopathy, present in > 90% of Alzheimer's disease brains and a major cause of intracerebral haemorrhage in the elderly; [70,87,176]), emerging roles in physiology (e.g., immune surveillance, central waste clearance and the proposed 'glymphatic' concepts; [95,121,124,137]), and as a critical gatekeeper for widespread central drug delivery [137]. Historical considerations: CSF/ISF exchange and flow along low-resistance pathways including the PVS It was recognized over one century ago that CSF was primarily formed by the choroid plexuses in the ventricles of the brain, through which it flowed before reaching the cranial and spinal SAS [38,186].…”

“…Filled with a fluid in potential communication with both CSF and ISF but also containing connective tissue, scattered cells, and basement membranes (BM), the PVS is a real compartment within the vascular connective tissue space of the adventitia [63,97,108,137,158,194,200] and possibly also the tunica media BM of large vessels [24,137,189] as well as a potential compartment within the basal lamina of capillaries [74,91,137,148]. Potential routes of entry from the CSF into the PVS include specialized pores, termed stomata, recently demonstrated on the adventitial lining cells of leptomeningeal vessels in the SAS of the rat by scanning electron microscopy [137] (Fig. 1b), confirming earlier decades-old identification of such structures in cats [199]; similar pores may also exist on the pia [30,146], providing an additional route into the PVS via the subpial space (discussed in [137]).…”

“…The perivascular spaces (PVS) 1 of cerebral blood vessels have in recent years been the subject of increasing research focus as pathways for CSF/ISF exchange [1,74,89,91,95,112,137,138,162], but controversy exists over their precise role [58,77,96,161,162]. Indeed, the glial components (astrocyte foot processes) that provide the outer boundary of the PVS within the parenchyma have been proposed to serve a special function for CNS clearance and waste turnover, forming the basis for a so-called 'glymphatic' circulation [91,124] that may potentially allow a more complete exchange of CSF and ISF at both superficial and deep sites spanning the entire neural axis.…”