Widespread hyperexcitability of nociceptor somata outlasts enhanced avoidance behavior after incision injury.

Nociceptors with somata in dorsal root ganglia (DRGs) readily switch from an electrically silent state to a hyperactive state of tonic, nonaccommodating, low-frequency, irregular discharge of action potentials (APs). Spontaneous activity (SA) during this state is present in vivo in rats months after spinal cord injury (SCI) and has been causally linked to SCI pain. Intrinsically generated SA and, more generally, ongoing activity (OA) are induced by various neuropathic conditions in rats, mice, and humans and are retained in nociceptor somata after dissociation and culturing, providing a powerful tool for investigating its mechanisms and functions.

Induction of long-term hyperexcitability by memory-related cAMP signaling in isolated nociceptor cell bodies.

Persistent hyperactivity of nociceptors is known to contribute significantly to long-lasting sensitization and ongoing pain in many clinical conditions. It is often assumed that nociceptor hyperactivity is mainly driven by continuing stimulation from inflammatory mediators. We have tested an additional possibility: that persistent increases in excitability promoting hyperactivity can be induced by a prototypical cellular signaling pathway long known to induce late-phase long-term potentiation (LTP) of synapses in brain regions involved in memory formation.

Mechanism of gabapentinoid potentiation of opioid effects on cyclic AMP signaling in neuropathic pain.

Over half of spinal cord injury (SCI) patients develop opioid-resistant chronic neuropathic pain. Safer alternatives to opioids for treatment of neuropathic pain are gabapentinoids (e.g.

Induction of long-term hyperexcitability by memory-related cAMP signaling in isolated nociceptor cell bodies.

Persistent hyperactivity of nociceptors is known to contribute significantly to long-lasting sensitization and ongoing pain in many clinical conditions. It is often assumed that nociceptor hyperactivity is mainly driven by continuing stimulation from inflammatory mediators. We have tested an additional possibility: that persistent increases in excitability promoting hyperactivity can be induced by a prototypical cellular signaling pathway long known to induce late-phase long-term potentiation (LTP) of synapses in brain regions involved in memory formation.

Structure of adenylyl cyclase 5 in complex with Gβγ offers insights into ADCY5-related dyskinesia.

The nine different membrane-anchored adenylyl cyclase isoforms (AC1-9) in mammals are stimulated by the heterotrimeric G protein, Gα, but their response to Gβγ regulation is isoform specific. In the present study, we report cryo-electron microscope structures of ligand-free AC5 in complex with Gβγ and a dimeric form of AC5 that could be involved in its regulation. Gβγ binds to a coiled-coil domain that links the AC transmembrane region to its catalytic core as well as to a region (C) that is known to be a hub for isoform-specific regulation.

Stabilization of interdomain interactions in G protein α subunits as a determinant of Gα subtype signaling specificity.

Highly homologous members of the Gα family, Gα, have distinct tissue distributions and physiological functions, yet their biochemical and functional properties are very similar. We recently identified PDZ-RhoGEF (PRG) as a novel Gα effector that is poorly activated by Gα. In a proteomic proximity labeling screen we observed a strong preference for Gα relative to Gα with respect to engagement of a broad range of potential targets.

Widespread latent hyperactivity of nociceptors outlasts enhanced avoidance behavior following incision injury.

Nociceptors with somata in dorsal root ganglia (DRGs) exhibit an unusual readiness to switch from an electrically silent state to a hyperactive state of tonic, nonaccommodating, low-frequency, irregular discharge of action potentials (APs). Ongoing activity (OA) during this state is present in vivo in rats months after spinal cord injury (SCI), and has been causally linked to SCI pain. OA induced by various neuropathic conditions in rats, mice, and humans is retained in nociceptor somata after dissociation and culturing, providing a powerful tool for investigating its mechanisms and functions.

The Concise Guide to PHARMACOLOGY 2023/24: Enzymes.

The Concise Guide to PHARMACOLOGY 2023/24 is the sixth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of approximately 1800 drug targets, and about 6000 interactions with about 3900 ligands. There is an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.

Isoform Specific Regulation of Adenylyl Cyclase 5 by Gβγ.

The nine different membrane-anchored adenylyl cyclase isoforms (AC1-9) in mammals are stimulated by the heterotrimeric G protein Gα, but their response to Gβγ regulation is isoform-specific. For example, AC5 is conditionally activated by Gβγ. Here, we report cryo-EM structures of ligand-free AC5 in complex with Gβγ and of a dimeric form of AC5 that could be involved in its regulation.

Stabilization of Interdomain Interactions in G protein α Subunits Determines Gα Subtype Signaling Specificity.

Highly homologous members of the Gα family, Gα, have distinct tissue distributions and physiological functions, yet the functional properties of these proteins with respect to GDP/GTP binding and regulation of adenylate cyclase are very similar. We recently identified PDZ-RhoGEF (PRG) as a novel Gα effector, however, it is poorly activated by Gα. Here, in a proteomic proximity labeling screen we observed a strong preference for Gα relative to Gα with respect to engagement of a broad range of potential targets.

POPDC1 scaffolds a complex of adenylyl cyclase 9 and the potassium channel TREK-1 in heart.

The establishment of macromolecular complexes by scaffolding proteins is key to the local production of cAMP by anchored adenylyl cyclase (AC) and the subsequent cAMP signaling necessary for cardiac functions. We identify a novel AC scaffold, the Popeye domain-containing (POPDC) protein. The POPDC family of proteins is important for cardiac pacemaking and conduction, due in part to their cAMP-dependent binding and regulation of TREK-1 potassium channels.

Major Differences in Transcriptional Alterations in Dorsal Root Ganglia Between Spinal Cord Injury and Peripheral Neuropathic Pain Models.

Chronic, often intractable, pain is caused by neuropathic conditions such as traumatic peripheral nerve injury (PNI) and spinal cord injury (SCI). These conditions are associated with alterations in gene and protein expression correlated with functional changes in somatosensory neurons having cell bodies in dorsal root ganglia (DRGs). Most studies of DRG transcriptional alterations have utilized PNI models where axotomy-induced changes important for neural regeneration may overshadow changes that drive neuropathic pain.

Macrophage Migration Inhibitory Factor (MIF) Makes Complex Contributions to Pain-Related Hyperactivity of Nociceptors after Spinal Cord Injury.

Neuropathic pain is a major, inadequately treated challenge for people with spinal cord injury (SCI). While SCI pain mechanisms are often assumed to be in the CNS, rodent studies have revealed mechanistic contributions from primary nociceptors. These neurons become chronically hyperexcitable after SCI, generating ongoing electrical activity that promotes ongoing pain.

Structural basis of adenylyl cyclase 9 activation.

Adenylyl cyclase 9 (AC9) is a membrane-bound enzyme that converts ATP into cAMP. The enzyme is weakly activated by forskolin, fully activated by the G protein Gαs subunit and is autoinhibited by the AC9 C-terminus. Although our recent structural studies of the AC9-Gαs complex provided the framework for understanding AC9 autoinhibition, the conformational changes that AC9 undergoes in response to activator binding remains poorly understood.

Physiological roles of mammalian transmembrane adenylyl cyclase isoforms.

Adenylyl cyclases (ACs) catalyze the conversion of ATP to the ubiquitous second messenger cAMP. Mammals possess nine isoforms of transmembrane ACs, dubbed AC1-9, that serve as major effector enzymes of G protein-coupled receptors (GPCRs). The transmembrane ACs display varying expression patterns across tissues, giving the potential for them to have a wide array of physiological roles.

THE CONCISE GUIDE TO PHARMACOLOGY 2021/22: Enzymes.

The Concise Guide to PHARMACOLOGY 2021/22 is the fifth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of nearly 1900 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.

G protein-coupled receptor-effector macromolecular membrane assemblies (GEMMAs).

G protein-coupled receptors (GPCRs) are the largest group of receptors involved in cellular signaling across the plasma membrane and a major class of drug targets. The canonical model for GPCR signaling involves three components - the GPCR, a heterotrimeric G protein and a proximal plasma membrane effector - that have been generally thought to be freely mobile molecules able to interact by 'collision coupling'. Here, we synthesize evidence that supports the existence of GPCR-effector macromolecular membrane assemblies (GEMMAs) comprised of specific GPCRs, G proteins, plasma membrane effector molecules and other associated transmembrane proteins that are pre-assembled prior to receptor activation by agonists, which then leads to subsequent rearrangement of the GEMMA components.

Serotonin enhances depolarizing spontaneous fluctuations, excitability, and ongoing activity in isolated rat DRG neurons via 5-HT receptors and cAMP-dependent mechanisms.

Ongoing activity in nociceptors, a driver of spontaneous pain, can be generated in dorsal root ganglion neurons in the absence of sensory generator potentials if one or more of three neurophysiological alterations occur - prolonged depolarization of resting membrane potential (RMP), hyperpolarization of action potential (AP) threshold, and/or increased amplitude of depolarizing spontaneous fluctuations of membrane potential (DSFs) to bridge the gap between RMP and AP threshold. Previous work showed that acute, sustained exposure to serotonin (5-HT) hyperpolarized AP threshold and potentiated DSFs, leading to ongoing activity if a separate source of maintained depolarization was present. Cellular signaling pathways that increase DSF amplitude and promote ongoing activity acutely in nociceptors are not known for any neuromodulator.

Depolarization-Dependent C-Raf Signaling Promotes Hyperexcitability and Reduces Opioid Sensitivity of Isolated Nociceptors after Spinal Cord Injury.

Chronic pain caused by spinal cord injury (SCI) is notoriously resistant to treatment, particularly by opioids. After SCI, DRG neurons show hyperactivity and chronic depolarization of resting membrane potential (RMP) that is maintained by cAMP signaling through PKA and EPAC. Importantly, SCI also reduces the negative regulation by Gαi of adenylyl cyclase and its production of cAMP, independent of alterations in G protein-coupled receptors and/or G proteins.

G protein-regulated endocytic trafficking of adenylyl cyclase type 9.

GPCRs are increasingly recognized to initiate signaling via heterotrimeric G proteins as they move through the endocytic network, but little is known about how relevant G protein effectors are localized. Here we report selective trafficking of adenylyl cyclase type 9 (AC9) from the plasma membrane to endosomes while adenylyl cyclase type 1 (AC1) remains in the plasma membrane, and stimulation of AC9 trafficking by ligand-induced activation of Gs-coupled GPCRs. AC9 transits a similar, dynamin-dependent early endocytic pathway as ligand-activated GPCRs.

EPAC1 and EPAC2 promote nociceptor hyperactivity associated with chronic pain after spinal cord injury.

Chronic pain following spinal cord injury (SCI) is associated with electrical hyperactivity (spontaneous and evoked) in primary nociceptors. Cyclic adenosine monophosphate (cAMP) signaling is an important contributor to nociceptor excitability, and knockdown of the cAMP effector, exchange protein activated by cAMP (EPAC), has been shown to relieve pain-like responses in several chronic pain models. To examine potentially distinct roles of each EPAC isoform (EPAC1 and 2) in maintaining chronic pain, we used rat and mouse models of contusive spinal cord injury (SCI).

Nanometric targeting of type 9 adenylyl cyclase in heart.

Adenylyl cyclases (ACs) convert ATP into the classical second messenger cyclic adenosine monophosphate (cAMP). Cardiac ACs, specifically AC5, AC6, and AC9, regulate cAMP signaling controlling functional outcomes such as heart rate, contractility and relaxation, gene regulation, stress responses, and glucose and lipid metabolism. With so many distinct functional outcomes for a single second messenger, the cell creates local domains of cAMP signaling to correctly relay signals.

Identification of Novel Adenylyl Cyclase 5 (AC5) Signaling Networks in D and D Medium Spiny Neurons using Bimolecular Fluorescence Complementation Screening.

Adenylyl cyclase type 5 (AC5), as the principal isoform expressed in striatal medium spiny neurons (MSNs), is essential for the integration of both stimulatory and inhibitory midbrain signals that initiate from dopaminergic G protein-coupled receptor (GPCR) activation. The spatial and temporal control of cAMP signaling is dependent upon the composition of local regulatory protein networks. However, there is little understanding of how adenylyl cyclase protein interaction networks adapt to the multifarious pressures of integrating acute versus chronic and inhibitory vs.

Regulation of I Potassium Current by Isoproterenol in Adult Cardiomyocytes Requires Type 9 Adenylyl Cyclase.

The subunits KCNQ1 and KCNE1 generate the slowly activating, delayed rectifier potassium current, I, that responds to sympathetic stimulation and is critical for human cardiac repolarization. The A-kinase anchoring protein Yotiao facilitates macromolecular complex formation between I and protein kinase A (PKA) to regulate phosphorylation of KCNQ1 and I currents following beta-adrenergic stimulation. We have previously shown that adenylyl cyclase Type 9 (AC9) is associated with a KCNQ1-Yotiao-PKA complex and facilitates isoproterenol-stimulated phosphorylation of KCNQ1 in an immortalized cell line.