Predisposition of Neonatal Maternal Separation to Visceral Hypersensitivity via Downregulation of Small-Conductance Calcium-Activated Potassium Channel Subtype 2 (SK2) in Mice

Abstract

Background. Visceral hypersensitivity is a common occurrence of gastrointestinal diseases such as irritable bowel syndrome (IBS), wherein early-life stress (ELS) may have a high predisposition to the development of visceral hypersensitivity in adulthood, with the specific underlying mechanism still elusive. Herein, we assessed the potential effect of small-conductance calcium-activated potassium channel subtype 2 (SK2) in the spinal dorsal horn (DH) on the pathogenesis of visceral hypersensitivity induced by maternal separation (MS) in mice. Methods. Neonatal mice were subjected to the MS paradigm, an established ELS model. In adulthood, the visceral pain threshold and the abdominal withdrawal reflex (AWR) were measured with an inflatable balloon. The elevated plus maze, open field test, sucrose preference test, and forced swim test were employed to evaluate the anxiety- and depression-like behaviors. The expression levels of SK2 in the spinal DH were determined by immunofluorescence and western blotting. The mRNA of SK2 and membrane palmitoylated protein 2 (MPP2) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Electrophysiology was applied to evaluate the neuronal firing rates and SK2 channel-mediated afterhyperpolarization current (IAHP). The interaction between MPP2 and SK2 was validated by coimmunoprecipitation. Results. In contrast to the naïve mice, ethological findings in MS mice revealed lowered visceral pain threshold, more evident anxiety- and depression-like behaviors, and downregulated expression of membrane SK2 protein and MPP2 protein. Moreover, electrophysiological results indicated increased neuronal firing rates and decreased IAHP in the spinal DH neurons. Nonetheless, intrathecal injection of the SK2 channel activator 1-ethyl-2-benzimidazolinone (1-EBIO) in MS mice could reverse the electrophysiological alterations and elevate the visceral pain threshold. In the naïve mice, administration of the SK2 channel blocker apamin abated IAHP and elevated spontaneous neuronal firing rates in the spinal DH neurons, reducing the visceral pain threshold. Finally, disruption of the MPP2 expression by small interfering RNA (siRNA) could amplify visceral hypersensitivity in naïve mice. Conclusions. ELS-induced visceral pain and visceral hypersensitivity are associated with the underfunction of SK2 channels in the spinal DH.

Behaviors Related to Psychiatric Disorders and Pain Perception in C57BL/6J Mice During Different Phases of Estrous Cycle

“…Sucrose preference test (SPT) was performed as we previously reported (Wu et al, 2020). Briefly, mice were individually housed to habituate drinking water with two bottles fitted with ballpoint sippers for 2 days.…”

Early life stress induces irritable bowel syndrome from childhood to adulthood in mice

“…Early life is the critical window for the development of gut microbiota, gut and brain ( Laursen et al, 2021 ; Ratsika et al, 2021 ). Therefore, disturbance of this process, such as caused by ELS, may have a high predisposition to the development of IBS in adulthood ( Wu et al, 2020 ). We found that MS affected the composition of both ileal and fecal microbiota ( Figures 6 , 7 ), suggesting MS affected intestinal microbiota community from early life to adulthood.…”

“…Maternal separation (MS) is a classic animal model of IBS, which effectively mimics ELS ( Riba et al, 2018 ; Wong et al, 2019 ; Huang S. T. et al, 2021 ; Tao et al, 2022a ). Using MS model, some of the pathogeneses of IBS were widely studied, such as visceral hypersensitivity ( Wu et al, 2020 ; Huang S. T. et al, 2021 ; Wang et al, 2022 ; Tao et al, 2022a ), intestinal hyperpermeability ( Kuti et al, 2020 ; Torres-Maravilla et al, 2022 ), intestinal dysmotility ( Bülbül and Sinen, 2021 ), intestinal dysbiosis ( Rincel and Darnaudéry, 2020 ; Park et al, 2021 ), and anxiety-like and depressive-like behaviors ( Zhou et al, 2022 ; Favoretto et al, 2023 ). Also, Riba et al ( Riba et al, 2018 ) systematically studied the influence of MS on the function of the intestine, mimicking IBS’s main features, including intestinal hyperpermeability, visceral hypersensitivity, microbiota dysbiosis, bile acid malabsorption, and low grade inflammation in the intestine.…”

Neural circuits regulating visceral pain

“…Inhibition of SK2 channels increases neuronal excitability and enhances excitatory synaptic transmission, as indicated by an increased frequency of miniature excitatory postsynaptic currents and action potentials, leading to hyperexcitability and pain hypersensitivity 169 . Recent research has shown that visceral hypersensitivity is related to the downregulation of SK2 channel proteins as well as the inactivation of SK2 channels in PVN CRH neurons 15 , 170 , 171 . Injecting SK2 channel activators into the PVN can reverse visceral hypersensitivity.…”