Investigating matrix metalloproteinase 9 in excitation/inhibition balance and bloodbrain barrier function following early-life seizures in zebrafish

Date of Award

6-2025

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Jocelyn Lippman-Bell, Ph.D

Second Advisor

Michelle Kanther, Ph.D.

Third Advisor

Brian J. Balin, Ph.D.

Abstract

Prolonged early-life seizures (ELS) can be associated with a range of long-term neurological consequences, including the development of spontaneous, recurrent seizures or epilepsy later in life. The process through which epilepsy develops is called epileptogenesis. While many cellular and molecular changes have been described that could potentially play a role in post-ELS epileptogenesis, there is still no therapy that can prevent it. One molecule of interest is matrix metalloproteinase 9 (MMP-9), whose levels have been shown to increase significantly following seizure-induced tissue damage. MMP-9 is an enzyme involved in extracellular matrix remodeling, neuroinflammation, and synaptic plasticity, processes that are highly relevant to brain development and function. These MMP-9-related mechanisms may contribute to blood-brain barrier (BBB) disruption and excitation/inhibition (E/I) synaptic imbalance through maladaptive pathways triggered by early-life hyperexcitability. This study aimed to investigate whether ELS caused BBB disruption and/or E/I synaptic imbalance in zebrafish. It also examined whether inhibiting MMP-9 activity following ELS could mitigate these effects. We hypothesized that ELS would disrupt BBB integrity and E/I synaptic balance, and that MMP-9 inhibition would prevent these changes and restore balance between excitatory and inhibitory synaptic expression.

To test these hypotheses, early-life seizures (ELS) were induced in zebrafish larvae using the chemoconvulsant pentylenetetrazol (PTZ). To assess the role of MMP-9, some fish received post-ELS treatment with the MMP-9 inhibitor JNJ0966. Blood-brain barrier (BBB) integrity and synaptic balance were evaluated using tracer injections and immunohistochemistry at later developmental stages. Key findings demonstrated that ELS increased BBB permeability, suggesting a compromise in cerebral vascular integrity. Additionally, ELS altered synaptic expression, with increased excitatory synaptic markers and decreased inhibitory ones at 21 dpf, indicating a long-lasting disruption in the balance between excitation and inhibition. Treatment with the MMP-9 inhibitor JNJ0966 alleviated the excitatory synaptic changes, but did not restore BBB integrity or recover inhibitory synaptic expression. These results suggest that while MMP-9 inhibition can reduce some seizure-induced synaptic alterations, however, it may not fully restore BBB integrity or synaptic balance. This study provides evidence that ELS leads to long-term disruptions in BBB integrity and synaptic balance in zebrafish, potentially via MMP-9 activation. Further optimization of MMP-9 inhibition or exploration of alternative therapeutic targets may be needed to address these changes. These findings highlight the complexity of ELS-induced brain alterations and the potential of targeting MMP-9 as a therapeutic strategy for preventing long-term neurological consequences of early-life seizures.

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