8/8 NADIA U01 Long-Term Effects of Adolescent Alcohol on Pain
Research Component 8/8: Long-Term Effects of Adolescent Alcohol on Pain
Principal Investigators: Nicholas Gilpin, Ph.D., Louisiana State University Health, and Tiffany Wills, Ph.D., Louisiana State University Health
Adolescent alcohol use leads to persistent neural adaptations and behavioral dysregulation that increase the risk of developing alcohol use disorder (AUD). Acute alcohol reduces pain, and chronic pain (e.g., hyperalgesia) can promote alcohol drinking through negative reinforcing analgesic effects. Paradoxically, chronic alcohol produces hyperalgesia or worsen pre-existing pain states. Recently, we reported that medial central amygdala (CeA) projections to a midbrain region called the periaqueductal gray (vlPAG) are critical for mediating hyperalgesia in chronically alcohol exposed adult male rats. This chronic alcohol weakens synaptic connectivity between medial CeA and vlPAG in adult male rats, photostimulation of the CeA-vlPAG circuit rescues hyperalgesia in alcohol-dependent adult rats, and photoinhibition of this circuit produces hyperalgesia in naïve rats. Our lab and others find that antagonism of corticotropin-releasing factor type-1 receptors (CRFR1) in CeA reduces hyperalgesia associated with alcohol withdrawal, nicotine withdrawal and traumatic stress in adult rats. Our overarching hypotheses are that chronic intermittent alcohol exposure in adolescent rats (AIE) produces persist long-term effects on polymodal (i.e., mechanical and thermal) hyperalgesia that is mediated by weakened CeA-vlPAG connectivity and increased CRFR1 signaling in CeA. The CRFR1 signaling gating of CeA-vlPAG function is important for mediating AIE-induced hyperalgesia.
We include preliminary data showing that 1) AIE produces rapid and long-lasting thermal and mechanical hyperalgesia during adolescence and that this effect persists into adulthood (Fig. 1), 2) AIE reduces synaptic drive and excitatory/inhibitory ratio on vlPAG-projecting medial CeA neurons in adulthood (Fig. 3), 3) CRFR1 is expressed on vlPAG projecting cells (Fig. 2), and 4) validation data for a CRFR1:cre rat for CRFR1+ cell typespecific modulation of CeA outputs (Fig. 4). Because we also propose to challenge rats with a short-lasting inflammatory pain challenge in adulthood, we have also piloted dose-response effects of carrageenan on nociception in adult Wistar rats. Here, we propose aims that will test the hypotheses that AIE produces hyperalgesia during adolescence that persists into adulthood (Specific Aim 1), that AIE reduces synaptic drive and excitatory/inhibitory balance of synaptic transmission onto vlPAG-projecting CeA neurons via a CRFR1-dependent (Specific Aim 2), and that pharmacological, circuit-based, and epigenetic modulation of CRFR1+ PAG- projecting CeA neurons will rescue AIE-induced hyperalgesia and CeA-vlPAG circuit plasticity (Specific
Aim 3). Importantly, we propose specific collaborations with Research Component 6 (PI: Chandler) and 5 (PI: Crews) along with the Epigenetics Core (PI: Pandey) of the NADIA consortium. This proposal focuses on testing adolescent alcohol effects on pain-related outcomes and aligns with the overall goal of the NADIA consortium to examine the effects of adolescent alcohol exposure on the adult organism.
Adolescent alcohol use is a major contributor to the development of alcohol use disorder (AUD). Adolescent intermittent ethanol (AIE) vapor exposure is used in rats to model alcohol use in adolescent humans, and has profound circuit, cellular, molecular and genetic effects in the brain . Long after termination of adolescent alcohol exposure, adult rats exhibit increases in alcohol drinking, risk-taking behavior, sleep disruptions and anxiety-like behavior, all of which increase risks for developing AUD later in life. In humans, pain promotes alcohol drinking  and alcohol withdrawal worsens pain outcomes . Similarly, in adult rats, chronic alcohol produces hyperalgesia and worsens pre-existing pain [4,5]. Critical for this proposal, early life challenges predispose humans and animals to pain. In human children and adolescents, pain severity is positively associated with frequency of alcohol use and intoxication . Childhood trauma is associated with increased risk of chronic pain in adulthood relative to non-trauma controls , and childhood abuse is associated with more reported pain . Work in animals confirms this relationship by showing that early life traumatic stress induces hyperalgesia and nociceptor sensitization in adulthood . Collectively, the literature suggests that early life alcohol exposure and other challenges lead to persistent high-pain phenotypes. Here, we will combine these factors by testing the effect of adolescent alcohol exposure on pain-like phenotypes during adulthood. There is a gap in our understanding of how adolescent alcohol exposure alters nociception 1) during adolescent alcohol exposure, 2) in adults with a history of adolescent alcohol exposure, and 3) in response to a mild pain challenge during adulthood. Here, we address this gap by testing specific hypotheses about the neurobiological processes underlying adolescent alcohol effects on pain-like outcomes in adults with a history of adolescent alcohol exposure.
Recent work from our lab identified a critical role for central amygdala (CeA) projections to ventrolateral periaqueductal gray (vlPAG) in mediating alcohol withdrawal hyperalgesia in adult male rats; we showed that alcohol withdrawal reduced CeA-vlPAG connectivity, and this effect was mimicked by photoinhibition of CeAvlPAG neurons and blocked by photostimulation of CeA-vlPAG neurons . The mechanism for this weakened CeA-vlPAG connectivity is unknown, but uncovering this mechanism will be critical for identification of targets for reducing pain in individuals with a history of chronic alcohol use. Because it is known that alcohol withdrawal disrupts synaptic transmission via CRFR1-dependent mechanisms in CeA of adult rats [10,11], CRF-CRFR1 modulation of synaptic transmission in vlPAG-projecting CeA neurons may be a mechanism whereby adolescent alcohol alters pain-like outcomes later in life.
All prior work on alcohol-pain interactions has been performed in adult males. Our preliminary data show that AIE produces hyperalgesia that persists into adulthood in male rats, and also reduces the excitatory/inhibitory ratio and synaptic drive on vlPAG-projecting CeA cells from male rats. Here, we will test the effect of AIE on pain-like outcomes during adolescence and adulthood in male and female rats. It is critical to test females because men and women process pain differently, and animals exhibit sex differences in baseline nociception and responses to the anti-nociceptive effects of analgesic drugs [12-14].
Specific Aim 1 will test the hypotheses that AIE produces hyperalgesia that persists into adulthood, and that AIE increases the nociceptive response to low-dose carrageenan in adulthood. This aim will also measure AIE and carrageenan effects on epigenetic modifiers of CRF and CRFR1 genes in CeA of adult rats.
Specific Aim 2 will test the hypotheses that AIE decreases the excitatory/inhibitory balance of synaptic transmission onto vlPAG-projecting CeA neurons via a CRFR1-dependent mechanism, and that these changes persist into adulthood and are augmented by low-dose carrageenan in adulthood. Using slice electrophysiology, we will test AIE effects on intrinsic excitability & synaptic transmission in CeA-vlPAG neurons.
Specific Aim 3 will test the hypotheses that AIE-induced hyperalgesia is reversed by CRFR1 antagonism in CeA, epigenetic inhibition of CRF-CRFR1 signaling in CeA and activation of the CeA-vlPAG circuit. This aim will use Wistar rats and CRFR1:cre rats generated by the Gilpin Lab. We will use optogenetics and cre-dependent viruses for circuit experiments. In collaboration with the Epigenetics Core, we will use CRISPR-Cas9 technology to test the effect of CeA CRF and CRFR1 epigenetic regulators on AIE-induced hyperalgesia.
- Crews FT, Robinson DL, Chandler LJ, Ehlers CL, Mulholland PJ, Pandey SC, Rodd ZA, Spear LP, Swartzwelder HS, Vetreno RP (2019) Mechanisms of Persistent Neurobiological Changes Following Adolescent Alcohol Exposure: NADIA Consortium Findings. Alcohol Clin Exp Res 43:1806-1822.
- Riley 3rd JL, King C (2009) Self-report of alcohol use for pain in a multi-ethnic community sample. J Pain 10:944-52.
- Egli M, Koob GF, Edwards S (2012) Alcohol dependence as a chronic pain disorder. Neurosci Biobehav Rev 36:2179-92.
- Gatch MB, Lal H (1999) Effects of ethanol and ethanol withdrawal on nociception in rats. Alcohol Clin Exp Res 23:328-33.
- Avegno EM, Lobell TD, Itoga CA, Baynes BB, Whitaker AM, Weera MM, Edwards S, Middleton JW, Gilpin NW (2018) Central Amygdala Circuits Mediate Hyperalgesia in Alcohol-Dependent Rats. J Neurosci 38:7761-7773.
- Horn-Hofmann C, Trost Z, Hublet A, Mrug S, Van Damme J, Vervoort T (2018) The Relationship Between Pain Severity and Alcohol Use Among School-Aged Children and Adolescents: The Moderating Role of Drinking Motives. Pain Med 19:284-296.
- Davis DA, Luecken LJ, Zautra AJ (2005) Are reports of childhood abuse related to the experience of chronic pain in adulthood? A meta-analytic review of the literature. Clin J Pain 21:398-405.
- Sachs-Ericsson N, Kendall-Tackett K, Hernandez A (2007) Childhood abuse, chronic pain, and depression in the National Comorbidity Survey. Child Abuse Negl 31:531-547.
- Green PG, Chen X, Alvarez P, Ferrari LF, Levine JD (2011) Early-life stress produces muscle hyperalgesia and nociceptor sensitization in the adult rat. Pain 152:2549-2556.
- Roberto M, Madamba SG, Stouffer DG, Parsons LH, Siggins GR (2004) Increased GABA release in the central amygdala of ethanol-dependent rats. J Neurosci 24:10159-10166.
- Roberto M, Cruz MT, Gilpin NW, Sabino V, Schweitzer P, Bajo M, Cottone P, Madamba SG, Stouffer DG, Zorrilla EP, Koob GF, Siggins GR, Parsons LH (2010) Corticotropin releasing factor-induced amygdala gammaaminobutyric Acid release plays a key role in alcohol dependence. Biol Psychiatry 67:831-839.
- Berkley, K.J. (1997) Sex differences in pain. Behav Brain Sci, 20, 371-380; discussion 435-513.
- Mogil JS, Bailey AL (2010) Sex and gender differences in pain and analgesia. Prog Brain Res 186:141-157.
- Hurley RW, Adams MC (2008) Sex, gender, and pain: an overview of a complex field. Anesth Analg 107:309-317.