Trans-segmental imaging in the spinal cord of behaving mice

Abstract

Spinal cord circuits play crucial roles in transmitting and gating cutaneous somatosensory modalities, such as pain, but the underlying activity patterns within and across spinal segments in behaving mice have remained elusive. To enable such measurements, we developed a wearable widefield macroscope with a 7.9 mm2 field of view, subcellular lateral resolution, 2.7 mm working distance, and <10 g overall weight. We show that highly localized painful mechanical stimuli evoke widespread, coordinated astrocyte excitation across multiple spinal segments.

A perspective on astrocyte regulation of neural circuit function and animal behavior

“…• Tamoxifen inducible version of Split-Cre excitation using separate groups of mice (Sekiguchi et al, 2016;Shekhtmeyster, Carey, et al, 2021). As expected, innocuous sensory stimuli of increasing intensity elevated the number and amplitude of responding excitatory neurons, suggesting that astrocytes' microdomain transients are driven primarily by synaptic activity.…”

“…For example, using quantitative sensory and motor assays together with in vivo calcium imaging, superficial (i.e., lamina I-II) dorsal horn astrocytes were found to respond to innocuous sensory stimuli (tail pinch) with asynchronous increases in the frequency but not amplitude or duration of their microdomain transients. In contrast, noxious stimuli evoked concerted calcium transients throughout the superficial astrocyte syncytium within and across spinal segments, both in the presence and absence of a flight response (Sekiguchi et al, 2016;Shekhtmeyster, Duarte, et al, 2021). Using the same standardized behavioral assays, local neural activity was measured and compared to astrocyte Hirrlinger, Scheller, et al, 2009;Jullien et al, 2003;Kim, Kolesnikov, et al, 2021 Split-Cre-ERT2…”

“…Recent studies have also indicated that spinal cord astrocytes are regionally heterogeneous (Kronschläger et al, 2021;Molofsky et al, 2014;Shekhtmeyster, Carey, et al, 2021), that this heterogeneity is functionally relevant (Kelley et al, 2018;Kohro et al, 2020) and associated with distinct spatiotemporal excitation patterns. For example, using translaminar imaging in behaving mice with chronically implanted microprisms, astrocytes in sensory and premotor areas of the spinal dorsal horn were found to respond to noxious mechanical stimuli and locomotion in a lamina-specific manner (Shekhtmeyster, Carey, et al, 2021). In contrast, Tac1-expressing neurons, involved in pain processing, responded to the same noxious stimuli with calcium spiking in sensory regions but did not respond to locomotion, suggesting that lamina-specific astrocyte activity depends on the Similarly, ventral astrocytes seem to play essential roles in controlling motor behavior.…”

“…To date, most spinal cord studies involving astrocytes have been performed in reduced preparations or anesthetized animals, mainly due to the technical difficulty of obtaining stable recordings from this CNS region during animal behavior, which typically results in large amplitude and nonlinear tissue displacements ( Nelson et al., 2019 ). The few studies that have been conducted in awake mice demonstrate that anesthesia powerfully suppresses spinal neuron and astrocyte activity and that sensorimotor-driven astrocyte excitation is complex and regionally heterogeneous ( Sekiguchi et al, 2016 ; Shekhtmeyster, Carey, et al, 2021 ; Shekhtmeyster, Duarte, et al, 2021 ). For example, using quantitative sensory and motor assays together with in vivo calcium imaging, superficial (i.e., lamina I–II) dorsal horn astrocytes were found to respond to innocuous sensory stimuli (tail pinch) with asynchronous increases in the frequency but not amplitude or duration of their microdomain transients.…”

“…Recent studies have also indicated that spinal cord astrocytes are regionally heterogeneous ( Kronschläger et al, 2021 ; Molofsky et al, 2014 ; Shekhtmeyster, Carey, et al, 2021 ), that this heterogeneity is functionally relevant ( Kelley et al, 2018 ; Kohro et al., 2020 ) and associated with distinct spatiotemporal excitation patterns. For example, using translaminar imaging in behaving mice with chronically implanted microprisms, astrocytes in sensory and premotor areas of the spinal dorsal horn were found to respond to noxious mechanical stimuli and locomotion in a lamina-specific manner ( Shekhtmeyster, Carey, et al, 2021 ). In contrast, Tac1-expressing neurons, involved in pain processing, responded to the same noxious stimuli with calcium spiking in sensory regions but did not respond to locomotion, suggesting that lamina-specific astrocyte activity depends on the neuronal cell type.…”