The Neurokinin-1 Receptor is Expressed with Gastrin-Releasing Peptide Receptor in Spinal Interneurons and Modulates Itch

Abstract

The neurokinin-1 receptor (NK1R, encoded by Tacr1) is expressed in spinal dorsal horn neurons and has been suggested to mediate itch. However, previous studies relied heavily on neurotoxic ablation of NK1R spinal neurons, which limited further dissection of their function in spinal itch circuitry. Thus, we leveraged a newly developed Tacr1 CreER mouse line to characterize the role of NK1R spinal neurons in itch. We show that pharmacological activation of spinal NK1R and chemogenetic activation of Tacr1 CreER spinal neurons increases itch behavior, whereas pharmacological inhibition of spinal NK1R suppresses itch behavior. We use fluorescence in situ hybridization to characterize the endogenous expression of Tacr1 throughout the superficial and deeper dorsal horn, as well as the lateral spinal nucleus. Retrograde labeling studies from the parabrachial nucleus show that less than 20% of superficial Tacr1 CreER dorsal horn neurons are spinal projection neurons, and thus the majority of Tacr1 CreER are local interneurons. We then use a combination of in situ hybridization and ex vivo two-photon Ca 2+ imaging of the spinal cord to establish that NK1R and the gastrin-releasing peptide receptor (GRPR) are coexpressed within a subpopulation of excitatory superficial dorsal horn neurons. These findings are the first to describe a role for NK1R interneurons in itch and extend our understanding of the complexities of spinal itch circuitry..

The Implications of Pruritogens in the Pathogenesis of Atopic Dermatitis

“…However, another group showed that SP activated MRGPCR on sensory neurons to induce itching in mice [149,150]. NK1R is expressed in the DRG, spinal horns, and brain areas responsible for itch signaling [151][152][153], and a recent study demonstrated that spinal NK1R neurons mediate itching in mice [152]. In line with the findings above, NK1R may have a central role in potentiating pruritus from the periphery to the spinothalamic tract and then to the brain cortex.…”

“…Significant amounts of sensory neurons express SP or GRP [151]. Additionally, GRP is a crucial spinal neurotransmitter for itch [170], and the latest study showed that spinal dorsal horn NK1R neurons, which are responsible for spinal pruritus, contained GRPR [152], and upregulation of serum neurotensin levels and gene expression in the lesioned skin of patients with AD. The possible interaction between neurotensin and MCs may contribute to the pathogenesis of AD [171].…”

The Spino-Parabrachial Pathway for Itch

“…However, chemogenetic and optogenetic stimulation of Tacr1 spinal projection neurons (Deng et al, 2020 ; Barik et al, 2021 ) drive spontaneous pain-induced protective behaviors and suppress itch (Barik et al, 2021 ). In an independent study, chemogenetic activation of Tacr1 neurons promoted itch and lacked spontaneous pain behaviors (Sheahan et al, 2020 ). This contradiction can be explained due to the usage of different genetic strategies to target the Tacr1 population.…”

Intraspinal injection of adeno-associated viruses into the adult mouse spinal cord

“…A critical factor for successful delivery and expression of fluorescent proteins is the serotype of virus. In spinal cord research, anterogradely transporting AAVs containing the genome of serotype 2 packaged in capsids from serotypes 1, 2, 5, 8, or 9 (AAV2/1, AAV2/2, AAV2/5, AAV2/8, AAV2/9) have been routinely used for the manipulation of spinal neurons or ascending pathways ( Dougherty et al., 2013 ; Fink et al., 2014 ; Bourane et al., 2015 ; Foster et al., 2015 ; Ruder, Takeoka and Arber, 2016 ; Choi et al., 2020 ; Sheahan et al., 2020 ; Barik et al., 2021 ; Gatto et al., 2021 ) ( Figures 1 A and 1B), while AAV retro ( Tervo et al., 2016 ), which is a capsid variant of AAV2, is used for targeting descending neurons ( Esposito et al., 2014 ; Basaldella et al., 2015 ; Murray et al., 2018 ; Sathyamurthy et al., 2020 ; Usseglio et al., 2020 ) ( Figure 1 C). For more information on AAVs, please refer to the following reviews ( Li et al., 2020 ; Naso et al., 2017 ; Samulski and Muzyczka, 2014 ; Wang et al., 2019 ).…”

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Figure 1 Intraspinal delivery of viral vectors presents a powerful approach for addressing basic questions regarding the organization and function of spinal circuits Genetically-defined groups of spinal neurons and the circuits that they are embedded in can be manipulated using two different groups of AAVs – anterogradely transporting (A and B) and retrogradely transporting AAVs (C). By injecting recombinant anterogradely transporting AAVs into different segments of the spinal cord in wildtype mice or genetic reporter lines, it is possible to acutely manipulate select spatially defined populations of spinal neurons, specifically in adulthood ( Dougherty et al., 2013 ; Fink et al., 2014 ; Bourane et al., 2015 ; Foster et al., 2015 ; Ruder, Takeoka and Arber, 2016 ; Choi et al., 2020 ; Sheahan et al., 2020 ; Barik et al., 2021 ; Gatto et al., 2021 ) (A). By combining spinal injections of anterogradely transporting AAVs with stereotactic injections of retrogradely transporting AAVs in brain regions that are targeted by ascending spinal neurons, and/or by the placement of optic fibers or drug cannulae in specific brain regions, it is possible to manipulate ascending pathways in a circuit-specific manner ( Conner et al., 2021 ; Fink et al., 2014 ; Bouvier et al., 2015 ; François et al., 2017 ; Sheahan et al., 2020 ) (B).

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