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Highlights • Calcium imaging is an efficient way to dissect the activity of neurons in vivo. • GCaMP indicators can be expressed in specific cell populations for in vivo imaging. • Pain research have benefitted greatly from these features in the recent decade. • Preclinical research is shifting towards the analysis of pain models and mechanisms. • In vivo calcium imaging is the ideal tool for an efficient drug discovery paradigm., Somatosensation and pain are complex phenomena involving a rangeofspecialised cell types forming different circuits within the peripheral and central nervous systems. In recent decades, advances in the investigation of these networks, as well as their function in sensation, resulted from the constant evolution of electrophysiology and imaging techniques to allow the observation of cellular activity at the population level both in vitro and in vivo. Genetically encoded indicators of neuronal activity, combined with recent advances in DNA engineering and modern microscopy, offer powerful tools to dissect and visualise the activity of specific neuronal subpopulations with high spatial and temporal resolution. In recent years various groups developed in vivo imaging techniques to image calcium transients in the dorsal root ganglia, the spinal cord and the brain of anesthetised and awake, behaving animals to address fundamental questions in both the physiology and pathophysiology of somatosensation and pain. This approach, besides giving unprecedented details on the circuitry of innocuous and painful sensation, can be a very powerful tool for pharmacological research, from the characterisation of new potential drugs to the discovery of new, druggable targets within specific neuronal subpopulations. Here we summarise recent developments in calcium imaging for pain research, discuss technical challenges and advances, and examine the potential positive impact of this technique in early preclinical phases of the analgesic drug discovery process.

Aldosterone produces rapid, non-genomic, inhibition of basolateral intermediate conductance K(+) (IK(Ca)) channels in human colonic crypt cells but the intracellular second messengers involved are unclear. We therefore evaluated the role of protein kinase C (PKC) in aldosterone's non-genomic inhibitory effect on basolateral IK(Ca) channels in crypt cells from normal human sigmoid colon. Patch clamp studies revealed that in cell-attached patches, IK(Ca) channel activity decreased progressively to 38+/-8% (P

Understanding how excitability of sensory neurons is regulated is an important goal since this excitability underlies pain transmission and unfortunately almost everyone will suffer from inflammatory pain at some point in their life. Recent studies have identified expression of M-type K+ channels (encoded by KCNQ genes) in damage-sensing (nociceptive) sensory neurons, where they are thought to control excitability. Accordingly, receptor-induced inhibition of M-current in these neurons has been shown to contribute to peripheral sensitisation and inflammatory pain. Here we describe a new mechanism for downregulation of KCNQ channel expression in nociceptors. We identified binding sites for the transcriptional repressor REST within both the KCNQ2 and KCNQ3 genes. Dorsal root ganglia (DRG) neurons cultured in the presence of a REST-expressing adenovirus showed 7.39±0.11 fold (p≤0.05) increased REST protein, which led to a concomitant 2.20±0.09 fold (p≤0.05) decrease in KCNQ2 protein and a corresponding 7.65±0.49 fold (p≤0.01) reduction in M-current in DRG neurons, compared to vehicle control. We further show that REST protein expression was increased 3.65±0.80 fold in cultured DRG neurons in response to inflammatory stimulation (1 μM bradykinin, 1 μM histamine, 1 μM ATP, 10 μM PAR2-AP and 1 μM substance P for 48 hrs). Increases in REST correlated with a 1.76±0.38 (p≤0.05) fold decrease in KCNQ2 immunoreactivity. Similarly we observed a significant increase in REST mRNA (2.11±0.01 fold) and protein levels and a reciprocal downregulation of KCNQ2 (1.75±0.07 fold) and KCNQ3 (1.43±0.01 fold) transcripts in DRGs from animals with neuropathic nerve injury (partial sciatic nerve lesion, PSNL). We propose that transcriptional regulation of KCNQ channels by REST will have profound effects on neuronal excitability and may contribute to the mechanisms of peripheral sensitisation in chronic pain.