Spatiotemporal and anatomical analyses of P2X receptor-mediated neuronal and glial processing of sensory signals in the rat dorsal horn

Abstract

Extracellularly released adenosine triphosphate (ATP) modulates sensory signaling in the spinal cord. We analyzed the spatiotemporal profiles of P2X receptor-mediated neuronal and glial processing of sensory signals and the distribution of P2X receptor subunits in the rat dorsal horn. Voltage imaging of spinal cord slices revealed that extracellularly applied ATP (5-500 μM), which was degraded to adenosine and acting on P1 receptors, inhibited depolarizing signals and that it also enhanced long-lasting slow depolarization, which was potentiated after ATP was washed out. This post-ATP rebound potentiation was mediated by P2X receptors and was more prominent in the deep than in the superficial layer. Patch clamp recording of neurons in the superficial layer revealed long-lasting enhancement of depolarization by ATP through P2X receptors during the slow repolarization phase at a single neuron level. This depolarization pattern was different from that in voltage imaging, which reflects both neuronal and glial activities. By immunohistochemistry, P2X(1) and P2X(3) subunits were detected in neuropils in the superficial layer. The P2X(5) subunit was found in neuronal somata. The P2X(6) subunit was widely expressed in neuropils in the whole gray matter except for the dorsal superficial layer. Astrocytes expressed the P2X(7) subunit. These findings indicate that extracellular ATP is degraded into adenosine and prevents overexcitation of the sensory system, and that ATP acts on pre- and partly on postsynaptic neuronal P2X receptors and enhances synaptic transmission, predominantly in the deep layer. Astrocytes are involved in sensitization of sensory network activity more importantly in the superficial than in the deep layer.

Over-expression of P2X7 receptors in spinal glial cells contributes to the development of chronic postsurgical pain induced by skin/muscle incision and retraction (SMIR) in rats

“…In the present study, we found that P2X7Rs were also located mainly in microglia, to a lesser extent in astrocytes in the spinal dorsal horn after SMIR. This is also in line with previous studies that apart from microglia, P2X7Rs are expressed in astrocytes of the cerebral cortex (Oliveira et al, 2011) and of the spinal dorsal horn (Aoyama et al, 2011). P2X7Rs play a pivotal role in the cross-talk between microglia and neurons (Sperlagh et al, 2006).…”

The Role of Purinergic Receptors in Cancer-Induced Bone Pain

“…Attempting to clarify the matter, Aoyama et al recently reported a systematic analysis of the distribution of all seven P2X receptors in the dorsal horn and compared their findings to earlier reports [46]. From this they concluded that P2X1 and P2X3 receptor subunits are densely distributed mainly in laminae I and II of the dorsal horn and are presumably expressed at the afferent nerve terminals [38].…”

Standardization of Size, Shape and Internal Structure of Spinal Cord Images: Comparison of Three Transformation Methods

“…VSD imaging techniques have been applied, e.g., to the sensory system of the spinal cord [17]–[19] as well as to the respiratory central pattern generator of the brainstem [20]–[23]. Also, mathematical methods to more efficiently detect neural activities from spatio-temporal data obtained by VSD imaging have been developed [23], [24].…”

“…The procedures using animals were done in accordance with the NIH Guide for the Care and Use of Laboratory Animals and were approved by the Animal Research Committee of Shimane University School of Medicine. The histological technique of NeuN staining has been described elsewhere [17]. Briefly, neonatal rats (Wistar, 4–5 day-old, n = 4) were deeply anesthetized by intraperitoneal injection of chloral hydrate (350 mg/kg), perfused transcardially with saline, and fixed with 4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.3).…”

“…Some standardization methods have been suggested not merely for brain but also for the liver or breast [13]–[16]. In animal studies, fluorescent voltage-sensitive dye (VSD) imaging has been widely applied to analyze spatio-temporal patterns of neural electric activities in the brain and spinal cord [17]–[24]. However, neither a registration nor standardization method for VSD imaging has ever been proposed for the brain or spinal cord of animals.…”