Doctor of Philosophy
Physiology and Pharmacology
Dr. Michael O. Poulter
Stress increases the frequency by which epileptic seizures occur. Corticotropin-releasing factor (CRF) coordinates neuroendocrine, autonomic and behavioral response to stress. This thesis sought to study the cellular and molecular mechanisms by which CRF regulates the activity of neural circuits in the piriform cortex (PC) in normal and epileptic states. The PC is richly innervated by CRF and 5-HT containing axons arising from the central amygdala and raphe nucleus. CRFR1 and 5-HT2A/CRs have been shown to interact in a manner where CRFR activation subsequently potentiates the activity of 5-HT2A/CRs. The first purpose of this thesis was to determine how the activation of CRFR1 and/or 5-HT2Rs modulates PC activity at both the circuit and cellular level. Voltage-sensitive dye imaging showed that CRF acting through CRFR1 dampened activation of layer II in the PC and interneurons of the endopiriform nucleus. Application of the selective 5-HT2A/CRagonist 2,5-dimethoxy-4-iodoamphetamine (DOI) following CRFR1 activation potentiated this effect. Blocking the interaction between CRFR1 and 5-HT2R with a Tat-CRFR1-CT peptide abolished this potentiation. Application of forskolin did not mimic CRFR1 activity but instead blocked it, while a protein kinase A antagonist had no effect. However, activation and antagonism of protein kinase C (PKC) either mimicked or blocked CRF modulation respectively. DOI had no effect when applied alone indicating that the prior activation of CRFR1 receptors was critical for the DOI activity. This data shows that CRF and 5-HT, acting through 5-HT2A/CRs, reduce the activation of the PC. This modulation may be an important blunting mechanism of stressor behaviors mediated through the olfactory cortex.
Anxiety and stress conditions induce neurons arising from the central amygdala and local interneurons to release CRF in PC, where it normally dampens excitability. The second aim
of this thesis was to determine the role of CRF in stress associated epilepsy. We showed that CRF increased the excitability of PC in rats subjected to kindling, a model of temporal lobe epilepsy. In non-kindled rats, CRF activates its receptor, a G protein-coupled receptor (GPCR) and signals through a Gaq/11 mediated pathway as identified in the first aim of this thesis. After seizure induction, CRF signaling occurred through a pathway involving Gas. This change in signaling was associated with reduced abundance of regulator of G-protein signaling protein-2 (RGS2), which promotes the switch in CRFR1 signaling cascade to a Gas dependent mechanism. RGS2 knockout mice responded to CRF in a similar manner as epileptic rats. These observations indicate that seizures produce changes in neuronal signaling that can increase seizure occurrence by converting a beneficial stress response into an epileptic trigger.
People with traumatic brain injury often develop epileptic seizures. The mechanisms underlying this are poorly understood. Considerable evidence suggests that association of stressful life experiences in brain injured patients lead them to develop post-traumatic stress disorder. CRF release in brain regions that are implicated in epileptogenesis make these situations worse. The third aim of this thesis was to understand the role of CRF in inducing excitability in PC after brain injury. We found that CRF has variable effects on the interneurons of ipsilateral and contralateral PC. Altogether, its actions lead to increased excitability of PC compared to healthy rat PC. The extent of excitability produced by CRF and the signaling mechanism of CRFR1 after brain injury were similar to CRF actions and CRFR1 signaling mechanism in kindling induced epilepsy.
Overall, this thesis study provides the basic mechanisms by which certain forms of epilepsy, both stress and injury induced develops. It also points out the important discovery of this project that is, the capability of GPCRs to switch signaling cascades depending on the pathological condition of the brain.
Narla, V V Chakravarthi, "Corticotropin releasing factor receptor type 1 signaling in epilepsy and traumatic brain injury" (2016). Electronic Thesis and Dissertation Repository. 4218.