Voltage-gated sodium channels (VGSCs), classically known to play a central role in excitability and signalling in nerves and muscles, have also been found to be expressed in a range of ...'non-excitable' cells, including lymphocytes, fibroblasts and endothelia. VGSC abnormalities are associated with various diseases including epilepsy, long-QT syndrome 3, Brugada syndrome, sudden infant death syndrome and, more recently, various human cancers. Given their pivotal role in a wide range of physiological and pathophysiological processes, regulation of functional VGSC expression has been the subject of intense study. An emerging theme is post-translational regulation and macro-molecular complexing by protein-protein interactions and intracellular trafficking, leading to changes in functional VGSC expression in plasma membrane. This partially involves endoplasmic reticulum associated degradation and ubiquitin-proteasome system. Several proteins have been shown to associate with VGSCs. Here, we review the interactions involving VGSCs and the following proteins: p11, ankyrin, syntrophin, beta-subunit of VGSC, papin, ERM and Nedd4 proteins. Protein kinases A and C, as well as Ca(2+)-calmodulin dependent kinase II that have also been shown to regulate intracellular trafficking of VGSCs by changing the balance of externalization vs. internalization, and an effort is made to separate these effects from the short-term phosphorylation of mature proteins in plasma membrane. Two further modulatory mechanisms are reciprocal interactions with the cytoskeleton and, late-stage, activity-dependent regulation. Thus, the review gives an updated account of the range of post-translational molecular mechanisms regulating functional VGSC expression. However, many details of VGSC subtype-specific regulation and pathophysiological aspects remain unknown and these are highlighted throughout for completeness.
Pain and emotional distress have a reciprocal relation. The amygdala has been implicated in emotional processing. The central nucleus of the amygdala (CeA) receives nociceptive information from the ...dorsal horn of spinal cord and is responsible for the central plasticity in chronic pain. Neuropathic pain is a type of severe chronic pain and can be strongly influenced by emotional components. Plastic changes in the CeA may play a key role in the development or maintenance or both of neuropathic pain. We studied the expression levels of proteins in the CeA of spinal nerve transection (SNT) model rats. Total tissue lysate proteins were separated by two-dimensional-gel electrophoresis (2D-PAGE). Gels from different time points were compared using Progenesis SameSpot software, and the spots with Fold Change greater than 2 were excised for protein identification by mass spectrometry. We identified more than 50 cytosolic proteins as significantly altered in their expression levels in the CeA of SNT rats, and most of these changes have been validated at mRNA levels by qRT-PCR. We also identified more than 40 membrane proteins as notably up- or down-regulated in the CeA of SNT model rats relative to a control using stable isotope dimethyl labeling nano-LC-MS/MS based proteomics and found that one such protein, doublecortin (DCX), a microtubule-associated protein expressed by neuronal precursor cells during development, is specifically localized in the membrane fraction without changes in total amount of the protein. Immunohistochemistry showed that doublecortin is expressed in processes in the CeA of rats 7 and 21 days after SNT surgery, suggesting that doublecortin is one of the proteins that may contribute to the plastic changes, namely, redevelopment or rewiring of neural networks, in the CeA in the neuropathic pain model. These dysregulated proteins may play roles in reciprocal relationships between pain and psychological distress in the amygdala and contribute to central sensitization. Data are available via ProteomeXchange with identifier PXD017473.
The possible association of intracellular Ca
2+
with metastasis in human cancer cells is poorly understood. We have studied Ca
2+
signaling in human prostate and breast cancer cell lines of strongly ...versus weakly metastatic potential in a comparative approach. Intracellular free Ca
2+
was measured using a membrane-permeant fluorescent Ca
2+
-indicator dye (Fluo-4 AM) and confocal microscopy. Spontaneous Ca
2+
oscillations were observed in a proportion of strongly metastatic human prostate and breast cancer cells (PC-3M and MDA-MB-231, respectively). In contrast, no such oscillations were observed in weakly/non metastatic LNCaP and MCF-7 cells, although a rise in the resting Ca
2+
level could be induced by applying a high-K
+
solution. Various parameters of the oscillations depended on extracellular Ca
2+
and voltage-gated Na
+
channel activity. Treatment with either tetrodotoxin (a general blocker of voltage-gated Na
+
channels) or ranolazine (a blocker of the persistent component of the channel current) suppressed the Ca
2+
oscillations. It is concluded that the functional voltage-gated Na
+
channel expression in strongly metastatic cancer cells makes a significant contribution to generation of oscillatory intracellular Ca
2+
activity. Possible mechanisms and consequences of the Ca
2+
oscillations are discussed.
The trafficking of Na sub(V)1.8 Swanwick, Richard S; Pristera, Alessandro; Okuse, Kenji
Neuroscience letters,
12/2010, Letnik:
486, Številka:
2
Journal Article
Recenzirano
The a-subunit of tetrodotoxin-resistant voltage-gated sodium channel Na sub(V)1.8 is selectively expressed in sensory neurons. It has been reported that Na sub(V)1.8 is involved in the transmission ...of nociceptive information from sensory neurons to the central nervous system in nociceptive and neuropathic pain conditions. Thus Na sub(V)1.8 has been a promising target to treat chronic pain. Here we discuss the recent advances in the study of trafficking mechanism of Na sub(V)1.8. These pieces of information are particularly important as such trafficking machinery could be new targets for painkillers.
The trafficking of Na(V)1.8 Swanwick, Richard S; Pristerá, Alessandro; Okuse, Kenji
Neuroscience letters,
2010-Dec-10, 20101210, Letnik:
486, Številka:
2
Journal Article
Recenzirano
Odprti dostop
The α-subunit of tetrodotoxin-resistant voltage-gated sodium channel Na(V)1.8 is selectively expressed in sensory neurons. It has been reported that Na(V)1.8 is involved in the transmission of ...nociceptive information from sensory neurons to the central nervous system in nociceptive 1 and neuropathic 24 pain conditions. Thus Na(V)1.8 has been a promising target to treat chronic pain. Here we discuss the recent advances in the study of trafficking mechanism of Na(V)1.8. These pieces of information are particularly important as such trafficking machinery could be new targets for painkillers.
We have examined the distribution of the sensory neuron-specific Na + channel Na v 1.8 (SNS/PN3) in nociceptive and non-nociceptive dorsal root ganglion (DRG) neurons and whether its distribution is ...related
to neuronal membrane properties. Na v 1.8-like immunoreactivity (Na v 1.8-LI) was examined with an affinity purified polyclonal antiserum (SNS11) in rat DRG neurons that were classified according
to sensory receptive properties and by conduction velocity (CV) as C-, Aδ- or Aα/β. A significantly higher proportion of nociceptive
than low threshold mechanoreceptive (LTM) neurons showed Na v 1.8-LI, and nociceptive neurons had significantly more intense immunoreactivity in their somata than LTM neurons. Results
showed that 89, 93 and 60 % of C-, Aδ- and Aα/β-fibre nociceptive units respectively and 88 % of C-unresponsive units were
positive. C-unresponsive units had electrical membrane properties similar to C-nociceptors and were considered to be nociceptive-type
neurons. Weak positive Na v 1.8-LI was also present in some LTM units including a C LTM, all Aδ LTM units (D hair), about 10 % of cutaneous LTM Aα/β-units,
but no muscle spindle afferent units. Na v 1.8-LI intensity was negatively correlated with soma size (all neurons) and with dorsal root CVs in A- but not C-fibre neurons.
Na v 1.8-LI intensity was positively correlated with action potential (AP) duration (both rise and fall time) in A-fibre neurons
and with AP rise time only in positive C-fibre neurons. It was also positively correlated with AP overshoot in positive neurons.
Thus high levels of Na v 1.8 protein may contribute to the longer AP durations (especially in A-fibre neurons) and larger AP overshoots that are typical
of nociceptors.
K(V)1.1 is a Shaker homologue K(+) channel that contributes to the juxta-paranodal membrane conductance in myelinated axons, and is blocked by fampridine (4-aminopyridine), used to treat the symptoms ...of multiple sclerosis. The present experiments investigate K(V)1.1 function in primary sensory neurons and A-fibres, and help define its characteristics as a drug-target using sequence specific small-interfering RNAs (siRNAs). siRNA (71nM) was used to knock-down functional expression of K(V)1.1 in sensory neurons (>25μm in apparent diameter) in culture, and was also delivered intrathecally in vivo (9.3μg). K(+) channel knock-down in sensory neurons was found to make the voltage-threshold for action potential generation significantly more negative than in control (p=0.02), led to the breakdown of accommodation and promoted spontaneous action potential firing. Exposure to dendrotoxin-K (DTX-K, 10-100nM) also selectively abolished K(+) currents at negative potentials and made voltage-threshold more negative, consistent with K(V)1.1 controlling excitability close to the nominal resting potential of the neuron cell body, near -60mV. Introduction of one working siRNA sequence into the intrathecal space in vivo was associated with a small increase in the amplitude of the depolarising after-potential in sacral spinal roots (p<0.02), suggesting a reduction in the number of working K(+) channels in internodal axon membrane. Our study provides evidence that K(V)1.1 contributes to the control of peripheral sensory nerve excitability, and suggests that its characteristics as a putative drug target can be assessed by siRNA transfection in primary sensory neurons in vitro and in vivo.
PDF
