Date of Degree

9-2019

Document Type

Dissertation

Degree Name

Ph.D.

Program

Biology

Advisor

Dan P. McCloskey

Committee Members

Joshua Brumberg

Patricia Rockwell

Asohan Amarasingham

Sabina Hrabětová

Subject Categories

Biophysics | Cell Biology | Molecular and Cellular Neuroscience

Keywords

Naked mole-rat, extracellular space, extracellular matrix, diffusion, viability, ischemia

Abstract

Hyaluronan (HA; Hyaluronic Acid), a primary scaffolding component of the brain extracellular matrix, serves as an integral structural component to the brain extracellular space (ECS). The fossorial African naked mole-rat (Heterocephalus glaber; NM-R), a mammal which lives in a low-oxygen environment and is capable of tolerating hypoxia and hypercapnia, has been shown to synthesize and sustain a unique high-molecular-mass variant of hyaluronan macromolecule (HMM-HA). This body of work highlights HA’s role in mediating the interplay between brain ECM composition, ECS structure, and cell viability.

Here we employ the NM-R as a unique animal model to observe the role of the extracellular matrix in maintaining brain microstructure and regulating diffusion in the brain ECS in health and disease states, such as ischemia. This portion of the work has produced a number of key findings, notably: 1) NM-R brain ECM expressing HMM-HA increases the hindrance to diffusion of macromolecules, and cleavage of HA further enhances this hindrance; 2) The ECS microstructure of the NM-R is remarkably resilient to focal ischemic insult, resisting significant changes in its tortuosity measure; 3) Hypo-osmotic stress differentially alters the ECS microstructure of the NM-R brain, suggesting an alternative mechanism of maintaining tissue viability.

The size of HA has the potential of influencing cell behavior, notably viability. Thus, we also explore HA’s potential, based on various sizes, in conferring tolerance and enhanced cell viability in wide extracellular pH excursions. Tissue acid/base balance is compromised during prolonged hypoxia or an acute ischemic insult, shifting towards severe acidosis, and ultimately influencing cell viability. Key findings from this portion of work: 4) Titration experiments indicate that the presence of HA differentially alters buffering dynamics in solution depending on the molecular weight of HA. 5) Under neutral pH conditions, NM-R neuronal viability is enhanced in a concentration-dependent manner when a high-molecular-weight HA species is added exogenously to primary neuron cultures; 6) Acute manipulation of extracellular pH in slice and primary neuron cultures did not stimulate HA production on the time scale investigated.

Together, these findings highlight the contribution of a unique HA variant in the organization of the ECS microstructure and buffering against dramatic brain metabolic shifts. The primary goal of this research is to posit that ECM composition, and HA in particular, dynamically influences the intracellular and extracellular compartments of the brain microenvironment, and ultimately operates at the interface of brain physiology and external environment of the organism.

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