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Diabetic neuropathic pain (DNP) is a diabetic complication that causes severe pain and deeply impacts the quality of the sufferer’s daily life. Currently, contemporary clinical treatments for DNP generally exhibit a deficiency in effectiveness. Electroacupuncture (EA) is recognized as a highly effective and safe treatment for DNP with few side effects. Regrettably, the processes via which EA alleviates DNP are still poorly characterized. Transient receptor potential vanilloid 1 (TRPV1) and phosphorylated calcium/calmodulin-dependent protein kinase II (p-CaMKII) are overexpressed on spinal cord dorsal horn (SCDH) in DNP rats, and co-localization is observed between them. Capsazepine, a TRPV1 antagonist, effectively reduced nociceptive hypersensitivity and downregulated the overexpression of phosphorylated CaMKIIα in rats with DNP. Conversely, the CaMKII inhibitor KN-93 did not have any impact on TRPV1. EA alleviated heightened sensitivity to pain caused by nociceptive stimuli and downregulated the level of TRPV1, p-CaMKIIα, and phosphorylated cyclic adenosine monophosphate response element-binding protein (p-CREB) in DNP rats. Intrathecal injection of capsaicin, on the other hand, reversed the above effects of EA. These findings indicated that the CaMKII/CREB pathway on SCDH is located downstream of TRPV1 and is affected by TRPV1. EA alleviates DNP through the TRPV1-mediated CaMKII/CREB pathway. [ABSTRACT FROM AUTHOR]

The modulation of TRPV1 emerges as a promising strategy for dental pain management. This study aimed to assess TRPV1 modulation in a human odontoblast-like cell model using Capsazepine (CZP) loaded in a nanogel delivery system. Gelatin nanogels, synthesized via the emulsification-gelation technique, were characterized and loaded with the TRPV1 antagonist, CZP. HPLC determined a remarkable 67.5 ± 0.04% CZP loading efficiency, with 71.7% of nanogels falling within the 300–950 nm size range, as evidenced by light microscopy. Moreover, CZP-loaded nanogels had a low cytotoxicity. An FTIR analysis showed no adverse chemical interactions, ensuring stability and active release. When examining biological responses, TRPV1 expression and channel activity were assessed in odontoblast-like cells. On the fifth day post-treatment, cells treated with CZP-loaded nanogels exhibited an increased TRPV1 expression and a reduction in calcium fluxes after agonist stimulus (F/F0 ratio 1.18 ± 0.18), resembling the response in free CZP-treated cells (1.28 ± 0.15). A two-way analysis of variance and the Tukey's test were used to determine statistical significance (p < 0.05). This delivery system, proven to be economical and straightforward, holds promise for dental pain management and potential local use. Local administration minimizes systemic adverse effects, making it a practical solution for releasing molecules in the oral cavity. [ABSTRACT FROM AUTHOR]

The modulation of TRPV1 emerges as a promising strategy for dental pain management. This study aimed to assess TRPV1 modulation in a human odontoblast-like cell model using Capsazepine (CZP) loaded in a nanogel delivery system. Gelatin nanogels, synthesized via the emulsification-gelation technique, were characterized and loaded with the TRPV1 antagonist, CZP. HPLC determined a remarkable 67.5 ± 0.04% CZP loading efficiency, with 71.7% of nanogels falling within the 300–950 nm size range, as evidenced by light microscopy. Moreover, CZP-loaded nanogels had a low cytotoxicity. An FTIR analysis showed no adverse chemical interactions, ensuring stability and active release. When examining biological responses, TRPV1 expression and channel activity were assessed in odontoblast-like cells. On the fifth day post-treatment, cells treated with CZP-loaded nanogels exhibited an increased TRPV1 expression and a reduction in calcium fluxes after agonist stimulus (F/F0 ratio 1.18 ± 0.18), resembling the response in free CZP-treated cells (1.28 ± 0.15). A two-way analysis of variance and the Tukey’s test were used to determine statistical significance (p < 0.05). This delivery system, proven to be economical and straightforward, holds promise for dental pain management and potential local use. Local administration minimizes systemic adverse effects, making it a practical solution for releasing molecules in the oral cavity.

Transient receptor potential vanilloid member 1 (TRPV1) is a heat and capsaicin receptor that allows cations to permeate and cause pain. As the molecular basis for temperature sensing, the heat capacity (ΔCp) model [D. E. Clapham, C. Miller, Proc. Natl. Acad. Sci. U.S.A. 108, 19492–19497 (2011).] has been proposed and experimentally supported. Theoretically, heat capacity is proportional to a variance in enthalpy, presumably related to structural fluctuation; however, the fluctuation of TRPV1 has not been directly visualized. In this study, we directly visualized single-molecule structural fluctuations of the TRPV1 channels in a lipid bilayer with the ligands resiniferatoxin (agonist, 1,000 times hotter than capsaicin) and capsazepine (antagonist) by high-speed atomic force microscopy. We observed the structural fluctuations of TRPV1 in an apo state and found that RTX binding enhances structural fluctuations, while CPZ binding suppresses fluctuations. These ligand-dependent differences in structural fluctuation would play a key role in the gating of TRPV1. [ABSTRACT FROM AUTHOR]

TRAIL, a tumor necrosis factor-related apoptosis-inducing ligand is a member of the tumor necrosis factor (TNF) superfamily, which is a cytokine that has shown a particularly precise and selective affinity towards death receptors that are overexpressed in cancer cells. While leaving the normal cells intact and unharmed, due to this property it has been the ligand of choice for highly precise cancer chemotherapeutic delivery system development. On numerous occasions, TRAIL has been used singularly and in combination with other drugs. It was observed that TRAIL had a tendency to be susceptible in terms of the cancer cells developing resistance against it. So TRAIL monotherapy was a bit of a rough patch for the molecule to become successful in the chemotherapy universe, however the conjugations and synergistic actions of TRAIL opened up new horizons which are discussed in this review with specific interest on colorectal cancer (CRC). [ABSTRACT FROM AUTHOR]

Piperine, the principle pungent compound in black peppers, is known to activate the capsaicin receptor TRPV1 ion channel. How piperine interacts with the channel protein, however, remains unclear. Here we show that piperine binds to the same ligand-binding pocket as capsaicin but in different poses. There was no detectable detrimental effect when T551 and E571, two major sites known to form hydrogen bond with capsaicin, were mutated to a hydrophobic amino acid. Computational structural modeling suggested that piperine makes interactions with multiple amino acids within the ligand binding pocket, including T671 on the pore-forming S6 segment. Mutations of this residue could substantially reduce or even eliminate piperine-induced activation, confirming that T671 is an important site. Our results suggest that the bound piperine may directly interact with the pore-forming S6 segment to induce channel opening. These findings help to explain why piperine is a weak agonist, and may guide future efforts to develop novel pharmaceutical reagents targeting TRPV1.

[Display omitted] • Capsazepine (CPZ) is a synthetic inhibitor of capsaicin-induced TRPV1 channel activation. • Using patch-clamp recordings, we discovered that CPZ inhibited I h amplitude in a concentration-dependent manner and slowed its activation time course. • A modified Markovian model anticipated the CPZ-induced decrease in the voltage-dependent hysteresis of I h. • CPZ also suppressed I Na in GH3 cells without affecting its activation or inactivation time course. • These results suggest that CPZ's inhibitory effects on I h and I Na are direct and not reliant on vanilloid receptor binding. A synthetic inhibitor of capsaicin-induced TRPV1 channel activation is called capsazepine (CPZ). In this study, we aimed to explore the effects of CPZ on hyperpolarization-activated cationic current (I h) and voltage-gated Na + current (I Na) in pituitary tumor (GH 3) cells. Through patch-clamp recordings, we found that CPZ concentration-dependently inhibited I h amplitude and slowed its activation time course. The IC 50 and K D values were 3.1 and 3.16 μM, respectively. CPZ also shifted the steady-state activation curve of I h towards a more hyperpolarized potential. However, there was no change in the gating charge of the curve. A modified Markovian model predicted the CPZ-induced decrease in the voltage-dependent hysteresis of I h. CPZ suppressed I Na in GH 3 cells, without altering its activation or inactivation time course. Additionally, exposure to CPZ reduced spontaneous firing. These findings suggest that CPZ's inhibitory effects on I h and I Na are direct and not dependent on vanilloid receptor binding. This could provide light on an unidentified ionic mechanism influencing the membrane excitability of neurons and endocrine or neuroendocrine cells in vivo. [ABSTRACT FROM AUTHOR]

Transient receptor potential vanilloid 1 (TRPV1), a non-selective ligand-gated cation channel with high permeability for Ca2+, has received considerable attention as potential therapeutic target for the treatment of several disorders including pain, inflammation, and hyperlipidemia. In particular, TRPV1 regulates lipid metabolism by mechanisms that are not completely understood. Interestingly, TRPV1 and lipids regulate each other in a reciprocal and complex manner. This review surveyed the recent literature dealing with the role of TRPV1 in the hyperlipidemia-associated metabolic syndrome. Besides TRPV1 structure, molecular mechanisms underlying the regulatory effect of TRPV1 on lipid metabolism such as the involvement of uncoupling proteins (UCPs), ATP-binding cassette (ABC) transporters, peroxisome proliferation-activated receptors (PPAR), sterol responsive element binding protein (SREBP), and hypoxia have been discussed. Additionally, this review extends our understanding of the lipid-dependent modulation of TRPV1 activity through affecting both the gating and the expression of TRPV1. The regulatory role of different classes of lipids such as phosphatidylinositol (PI), cholesterol, estrogen, and oleoylethanolamide (OEA), on TRPV1 has also been addressed.

Antagonism of transient receptor potential vanniloid-1 (TRPV1) and desensitization of transient receptor potential ankyrin-1 (TRPA1) nociceptors alleviate inflammatory bowel diseases (IBD)-associated chronic pain. However, there is limited literature available about their role in regulating the mucosal layer, its interaction with host physiology, and luminal microbial community. The present study focuses on the effects' intra rectal administration of capsazepine (modulator of TRPA1/TRPV1 expressing peptidergic sensory neurons) on colonic mucus production and gut health. We performed histological analysis, gut permeability alteration, gene expression changes, metabolite profiling, and gut microbial abundance in the ileum, colon, and cecum content of these animals. Intra rectal administration of capsazepine modulates TRPA1/TRPV1-positive nociceptors (behavioral pain assays) and resulted in damaged mucosal lining, increased gut permeability, and altered transcriptional profile of genes for goblet cell markers, mucus regulation, immune response, and tight junction proteins. The damage to mucosal lining prevented its role in enterosyne (short chain fatty acids) actions. These results suggest that caution must be exercised before employing TRPA1/TRPV1 modulation as a therapeutic option to alleviate pain caused due to IBD. [ABSTRACT FROM AUTHOR]

Bipolar disorder (BD) is a multifactorial chronic and refractory disease characterized by manic, depressive, and mixed mood episodes. Although epidemiological, and pathophysiological studies demonstrated a strong correlation between bipolar disorder and oxidative stress, precise etiology is still missing. Recent studies suggested the possible role of transient receptor potential channels (TRP) in the BD but, current knowledge is limited. Therefore, the current study investigates the possible role of TRPV1 in the ouabain-induced model of BD. The model was created with intracerebroventricular single dose ouabain (10−3 M) administration. Animals were treated with capsaicin, capsazepine, and lithium for seven days. Mania and depressive-like states were investigated with open-field, sucrose preference, and elevated plus maze tests. Oxidative stress was assessed by measuring total antioxidant and oxidant states, spectrophotometrically. The phosphorylation Glycogen synthase kinase-3β (GSK-3β) evaluated by western blotting. Our results demonstrated that capsaicin dose-dependently inhibited the ouabain-induced hyperlocomotion and depression. Although capsazepine exacerbated behavioral impairment, it did not show a significant effect on the antioxidant and oxidant states, and the effects of capsazepine on behaviors were abolished by combination with capsaicin. Additionally, capsaicin potently prevented the ouabain-induced decrease in GSK-3β phosphorylation. In contrast, capsazepine potentiated ouabain-induced decrease in GSK-3β phosphorylation and combination with capsaicin, suppressed the effect of capsazepine on GSK-3β phosphorylation. The effects of TRPV1 activation on oxidative stress and mania-like behaviors in the ouabain-induced BD model might be regulated by GSK-3β phosphorylation. [ABSTRACT FROM AUTHOR]

Background: Sustained Hif1α (hypoxic-inducible factor-1 alpha) accumulation plays a central role in osteogenic transdifferentiation and subsequent calcification. Capsaicin, the potent agonist of TRPV1 (transient receptor potential vanilloid type 1), was found to mitigate hypoxic-related injury and reverse phenotypic switch of vascular smooth muscle cells. However, its role in arterial calcification and the underlying mechanisms remain unexplored.Methods: We used data from Multi-Ethnic Study of Atherosclerosis to examine the association of coronary artery calcification and chili consumption. Chronic kidney disease mice and high phosphate-induced vascular smooth muscle cells calcification models were established to investigate the anticalcification effect of capsaicin, evaluated by calcium deposition and changes in phenotype markers.Results: Chili consumption was negatively correlated with coronary artery calcification and conferred a smaller progression burden during follow-up. Capsaicin reduced calcium deposition and osteogenic transdifferentiation both in vivo and in vitro. Using siTRPV1 (small interfering RNA and the antagonist of TRPV1), the anticalcification effect of capsaicin was abrogated. Hif1α was increased in phosphate-treated vascular smooth muscle cells and its degradation was accelerated by capsaicin. Retaining Hif1α stability using cobalt chloride (CoCl2) or MG132 abolished the protective effect of capsaicin. We further identified an increased expression of SIRT6 (Sirtuin 6) in response to capsaicin and confirmed the physical interaction between SIRT6 and Hif1α. Acetylated Hif1α was decreased, whereas hydroxylated Hif1α was increased under capsaicin treatment. Using immunohistochemistry analysis, we observed increased SIRT6 and reduced Hif1α in both SIRT6 transgenic and capsaicin-treated chronic kidney disease mice.Conclusions: Capsaicin facilitates deacetylation and degradation of Hif1α by upregulating SIRT6, which inhibits osteogenic transdifferentiation and protects against arterial calcification. These data highlight a promising therapeutic target for the management of arterial calcification. [ABSTRACT FROM AUTHOR]

Although the pharmacological activity of capsaicin has been explained by its specific binding to transient receptor potential vanilloid type 1, the amphiphilic structure of capsaicin may enable it to act on lipid bilayers. From a mechanistic point of view, we investigated whether capsaicin and its antagonist capsazepine interact with biomimetic membranes, and how capsazepine influences the membrane effect of capsaicin. Liposomal phospholipid membranes and neuro-mimetic membranes were prepared with 1,2-dipalmitoylphosphatidylcholine and with 1-palmitoyl-2-oleoylphosphatidylcholine and sphingomyelin plus cholesterol, respectively. These membrane preparations were subjected to reactions with capsaicin and capsazepine at 0.5–250 μM, followed by measuring fluorescence polarization to determine the membrane interactivity to modify the fluidity of membranes. Both compounds acted on 1,2-dipalmitoylphosphatidylcholine bilayers and changed membrane fluidity. Capsaicin concentration-dependently interacted with neuro-mimetic membranes to increase their fluidity at low micromolar concentrations, whereas capsazepine inversely decreased the membrane fluidity. When used in combination, capsazepine inhibited the effect of capsaicin on neuro-mimetic membranes. In addition to the direct action on transmembrane ion channels, capsaicin and capsazepine share membrane interactivity, but capsazepine is likely to competitively antagonize capsaicin's interaction with neuro-mimetic membranes at pharmacokinetically-relevant concentrations. The structure-specific membrane interactivity may be partly responsible for the analgesic effect of capsaicin. [ABSTRACT FROM AUTHOR]

Antagonism of transient receptor potential vanniloid-1 (TRPV1) and desensitization of transient receptor potential ankyrin-1 (TRPA1) nociceptors alleviate inflammatory bowel diseases (IBD)-associated chronic pain. However, there is limited literature available about their role in regulating the mucosal layer, its interaction with host physiology, and luminal microbial community. The present study focuses on the effects’ intra rectal administration of capsazepine (modulator of TRPA1/TRPV1 expressing peptidergic sensory neurons) on colonic mucus production and gut health. We performed histological analysis, gut permeability alteration, gene expression changes, metabolite profiling, and gut microbial abundance in the ileum, colon, and cecum content of these animals. Intra rectal administration of capsazepine modulates TRPA1/TRPV1-positive nociceptors (behavioral pain assays) and resulted in damaged mucosal lining, increased gut permeability, and altered transcriptional profile of genes for goblet cell markers, mucus regulation, immune response, and tight junction proteins. The damage to mucosal lining prevented its role in enterosyne (short chain fatty acids) actions. These results suggest that caution must be exercised before employing TRPA1/TRPV1 modulation as a therapeutic option to alleviate pain caused due to IBD.

Gang Wang, Xinglin Wang, Qian Gao, Ning Wang, Ming Zhou Department of Rehabilitation Medicine, First Medical Center of Chinese PLA General Hospital, Beijing 100853, People’s Republic of ChinaCorrespondence: Xinglin WangDepartment of Rehabilitation Medicine, First Medical Center of Chinese PLA General Hospital, 28 Fuxing Road, Beijing 100853, People’s Republic of ChinaTel +86 10 6693 7555Email 301wxl@sina.comPurpose: This study was aimed to investigate the effects of heat-conduction dry needling therapy on chronic myofascial pain syndrome (CMPS) in rats and explore the analgesic mechanisms by measuring the mRNA expression of transient receptor potential cation channel subfamily V member 1 (TRPV1) in musculus gastrocnemius.Methods: Seventy-two rats were randomly divided into five groups. Group A was the control group containing eight normal rats. Group B was the model group where 16 rats developed CMPS in musculus gastrocnemius but received no treatments. Groups C, D, and E containing 16 CMPS rats in each group were the trial groups receiving heat-conduction dry needling therapy. In detail, rats in Group C received dry needling treatment at the heating temperature of 44°C. Rats in Group D received intramuscular injection of capsazepine before receiving dry needling treatment with the heating temperature at 44°C. Rats in Group E received dry needling treatment with the heating temperature at 40°C. The mRNA expression of TRPV1, protein kinase C (PKC), and interleukin (IL)-6 in the needle insertion points of musculus gastrocnemius was measured at 24 hrs and 7 days after dry needling therapy.Results: Compared with untreated CMPS rats, the mRNA expression of TRPV1, PKC, and interleukin-6 (IL-6) was increased in rats receiving dry needling therapy with the heating temperature at 44°C for 24 hrs. However, the mRNA expression of TRPV1, PKC, and IL-6 in CMPS rats receiving injection of TRPV1 antagonist capsazepine into musculus gastrocnemius followed by temperature-needling at 44°C was analogous to that in untreated CMPS rats.Conclusion: Dry needling therapy with needles heated to 44°C could impact the activity of TRPV1/PKC/IL-6 pathway.Keywords: dry needling, myofascial pain syndrome, transient receptor potential, heat, capsazepine

Although the pharmacological activity of capsaicin has been explained by its specific binding to transient receptor potential vanilloid type 1, the amphiphilic structure of capsaicin may enable it to act on lipid bilayers. From a mechanistic point of view, we investigated whether capsaicin and its antagonist capsazepine interact with biomimetic membranes, and how capsazepine influences the membrane effect of capsaicin. Liposomal phospholipid membranes and neuro-mimetic membranes were prepared with 1,2-dipalmitoylphosphatidylcholine and with 1-palmitoyl-2-oleoylphosphatidylcholine and sphingomyelin plus cholesterol, respectively. These membrane preparations were subjected to reactions with capsaicin and capsazepine at 0.5–250 μM, followed by measuring fluorescence polarization to determine the membrane interactivity to modify the fluidity of membranes. Both compounds acted on 1,2-dipalmitoylphosphatidylcholine bilayers and changed membrane fluidity. Capsaicin concentration-dependently interacted with neuro-mimetic membranes to increase their fluidity at low micromolar concentrations, whereas capsazepine inversely decreased the membrane fluidity. When used in combination, capsazepine inhibited the effect of capsaicin on neuro-mimetic membranes. In addition to the direct action on transmembrane ion channels, capsaicin and capsazepine share membrane interactivity, but capsazepine is likely to competitively antagonize capsaicin’s interaction with neuro-mimetic membranes at pharmacokinetically-relevant concentrations. The structure-specific membrane interactivity may be partly responsible for the analgesic effect of capsaicin.

The expression of transient receptor potential protein channels increases intensively colon and breast cancer cells. We aimed to reveal the role of 5-fluorouracil (5FU) and capecitabine along with Transient Receptor Potential Vanilloid 1 (TRPV1) channels in breast and colon cancer cells. Breast (MCF-7) and colon (Caco-2) cells were cultured and the study was planned as 7 main groups. Cells in the group were incubated with 5FU and capecitabine for 24 h and then incubated with TRPV1 channel antagonist capsazepine and stimulator capsaicin. The effects of medicines were investigated on molecular pathways of apoptosis. It was concluded that the administration of TRPV1 channel stimulator capsaicin in both cancer cells significantly increased the degree of intracellular Ca2+ levels and apoptosis levels compared to the control group whereas, the use of TRPV1 channel inhibitor capsazepine, significantly decreased the degree of apoptosis levels. The apoptotic effects of 5FU and capecitabine on both colon and breast cancer cells are directly related to TRPV1 channels and TRPV1 channels play an important role in the apoptosis. The apoptotic cell lines activity of capecitabine was higher in breast cancer cells, while that of 5FU was more pronounced in colon cancer cells. [ABSTRACT FROM AUTHOR]

Sodium influx carried out by ion channels is one of the main regulators of water-salt and volume balance in cells of blood origin. Previously, we described amiloride-insensitive ENaC-like channels in human myeloid leukemia K562 cells; the intracellular regulatory mechanisms of the channels are associated with actin cytoskeleton dynamics. Recently, an extracellular mechanism of ENaC-like channels activation in K562 cells by the action of serine protease trypsin has been revealed. The other extracellular pathways that modulate ENaC (epithelial Na+ channel) activity and sodium permeability in transformed blood cells are not yet fully investigated. Here, we study the action of capsazepine (CPZ), as δ-ENaC activator, on single channel activity in K562 cells in whole-cell patch clamp experiments. Addition of CPZ (2 μM) to the extracellular solution caused an activation of sodium channels with typical features; unitary conductance was 15.1 ± 0.8 pS. Amiloride derivative benzamil (50 μM) did not inhibit their activity. Unitary currents and conductance of CPZ-activated channels were higher in Na+-containing extracellular solution than in Li+, that is one of the main fingerprints of δ-ENaC. The results of RT-PCR analysis and immunofluorescence staining also confirmed the expression of δ-hENaC (as well as α-, β-, γ-ENaC) at the mRNA and protein level. These findings allow us to speculate that CPZ activates amiloride-insensitive ENaC-like channels that contain δ-ENaC in К562 cells. Our data reveal a novel extracellular mechanism for ENaC-like activation in human leukemia cells. • δ-ENaC (as well as α-, β- γ-ENaC) is expressed in human myeloid leukemia K562 cells. • Capsazepine activates ENaC-like channels containing δ-ENaC in K562 cells. • Novel extracellular mechanism for ENaC-like activation in leukemia cells was revealed. [ABSTRACT FROM AUTHOR]

Transient receptor potential vanilloid 1 (TRPV1) has been implicated in peripheral inflammation and is a mediator of the inflammatory response to various noxious stimuli. However, the interaction between TRPV1 and N-methyl-D-aspartate (NMDA) receptors in the regulation of inflammatory pain remains poorly understood. This study aimed to investigate the analgesic effects of intrathecal administration of capsazepine, a TRPV1 antagonist, on carrageenan-induced inflammatory pain in mice and to identify its interactions with NMDA receptors. Inflammatory pain was induced by intraplantar injection of 2% carrageenan in male ICR mice. To investigate the analgesic effects of capsazepine, pain-related behaviors were evaluated using von Frey filaments and a thermal stimulator placed on the hind paw. TRPV1 expression and NMDA receptor phosphorylation in the spinal cord and glutamate concentration in the spinal cord and serum were measured. Intrathecal treatment with capsazepine significantly attenuated carrageenan-induced mechanical allodynia and thermal hyperalgesia. Moreover, carrageenan-enhanced glutamate and phosphorylation of NMDA receptor subunit 2B in the spinal cord were suppressed by capsazepine administration. These results indicate that TRPV1 and NMDA receptors in the spinal cord are associated with inflammatory pain transmission, and inhibition of TRPV1 may reduce inflammatory pain via NMDA receptors.

Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit fatty-acid amide hydrolase (FAAH), the enzyme responsible for the metabolism of anandamide, an endocannabinoid. It has been suggested that the mechanisms of action of NSAIDs could be due to inhibition of cyclooxygenase (COX) and also to an increase in endocannabinoid concentrations. In a previous study we have demonstrated that the local analgesic interaction between anandamide and ibuprofen (a non-specific COX inhibitor) was synergistic for the acute and inflammatory phases of the formalin test. To test this hypothesis further, we repeated similar experiments with rofecoxib (a selective COX-2 inhibitor) and also measured the local concentrations of anandamide, and of two fatty-acid amides, oleoylethanolamide and palmitoylethanolamide. We established the ED(50) for anandamide (34.52 pmol+/-17.26) and rofecoxib (381.72 pmol+/-190.86) and showed that the analgesic effect of the combination was synergistic. We also found that paw tissue levels of anandamide, oleoylethanolamide and palmitoylethanolamide were significantly higher when anandamide was combined with NSAIDs and that this effect was greater with rofecoxib. In conclusion, local injection of anandamide or rofecoxib was antinociceptive in a test of acute and inflammatory pain and the combination of anandamide with rofecoxib was synergistic. Finally, locally injected anandamide with either NSAID (ibuprofen or rofecoxib) generates higher amount of fatty-acid ethanolamides. The exact comprehension of the mechanisms involved needs further investigation.

Process of inflammation and complex interactions between immune and cancer cells within tumor microenvironment are known to drive and shape the outcome of the neoplastic disease. Recent studies increasingly show that ion channels can be used as potential targets to modulate immune response and to treat inflammatory disorders and cancer. The action of both innate and adaptive immune cells is tightly regulated by ionic signals provided by a network of distinct ion channels. TRPV1 channel, known as a capsaicin receptor, was recently documented to be expressed on the cells of the immune system but also aberrantly expressed in the several tumor types. It is activated by heat, protons, proinflammatory cytokines, and associated with pain and inflammation. TRPV1 channel is not only involved in calcium signaling fundamental for many cellular processes but also takes part in cell-environment crosstalk influencing cell behavior. Furthermore, in several studies, activation of TRPV1 by capsaicin was associated with anti-cancer effects. Therefore, TRPV1 provides a potential link between the process of inflammation, cancer and immunity, and offers new treatment possibilities. Nevertheless, in many cases, results regarding TRPV1 are contradictory and need further refinement. In this review we present the summary of the data related to the role of TRPV1 channel in the process of inflammation, cancer and immunity, limitations of the studies, and directions for future research.

Background and objectives Hyperpolarization-activation cyclic nucleotide-gated (HCN) channels contribute to the effects of lidocaine. Capsazepine (CPZ), a competitive inhibitor of capsaicin of transient receptor potential vanilloid-1 channel, has also been found to inhibit HCN channel currents (Ih). This study was designed to investigate whether CPZ could prolong durations of lidocaine in regional anesthesia. Methods Mouse HCN1 and HCN2 channels were expressed in human embryonic kidney 293 (HEK 293) cells. The effect of CPZ on Ih was measured by whole-cell patch-clamping recording. Sciatic nerve block model in mice was used for the study in vivo. The mice were randomly divided into seven groups, respectively, receiving lidocaine, CPZ, ZD7288 (HCN channel blocker), CPZ + lidocaine, ZD7288 + lidocaine, ZD7288 + CPZ + lidocaine, forskolin (an activator of adenylyl cyclase) + CPZ + lidocaine. Regional anesthetic durations of lidocaine were determined. Voltage-gated sodium channel currents (INa) and Ih were recorded in dorsal root ganglion neurons of mice. The effects of CPZ on INa and Ih with or without Cyclic adenosine monophosphate (cAMP) were assessed. Isolated mice sciatic nerve was prepared to evaluate the effect of CPZ on the compound action potentials (CAP). Results Capsazepine non-selectively inhibited transfected mHCN1 and mHCN2 channel currents in HEK 293 cells. In sciatic nerve block in vivo, compared to lidocaine alone, adding CPZ extended the durations of lidocaine for noxious sensory block (35.1 ± 3.3 vs. 20.3 ± 1.7 min), tactile sensory block (25.5 ± 4.4 vs. 20.0 ± 3.7 min), thermal sensory block (39.6 ± 6.6 vs. 26.8 ± 5.5 min), and motor function block (28.6 ± 4.1 vs. 20.9 ± 4.2 min). Duration of thermal sensory block was longer in CPZ + lidocaine group than that of ZD7288 + lidocaine group (39.6 ± 6.6 vs. 33.4 ± 4.5 min). Forskolin reversed the prolongation by CPZ on lidocaine durations. CPZ or ZD7288 alone did not produce typical regional anesthetic effects. Increased intracellular concentration of cAMP reversed the inhibition of CPZ on Ih. Although CPZ alone inhibited INa at the concentration more than 30 μM, it did not inhibit the CAP amplitudes in isolated sciatic nerves. CPZ dose-dependently enhanced the inhibitory effect of 1% lidocaine on the CAP amplitudes. Conclusions Capsazepine may prolong durations of lidocaine in peripheral nerve block by modulation of HCN channel currents.

Process of inflammation and complex interactions between immune and cancer cells within tumor microenvironment are known to drive and shape the outcome of the neoplastic disease. Recent studies increasingly show that ion channels can be used as potential targets to modulate immune response and to treat inflammatory disorders and cancer. The action of both innate and adaptive immune cells is tightly regulated by ionic signals provided by a network of distinct ion channels. TRPV1 channel, known as a capsaicin receptor, was recently documented to be expressed on the cells of the immune system but also aberrantly expressed in the several tumor types. It is activated by heat, protons, proinflammatory cytokines, and associated with pain and inflammation. TRPV1 channel is not only involved in calcium signaling fundamental for many cellular processes but also takes part in cell-environment crosstalk influencing cell behavior. Furthermore, in several studies, activation of TRPV1 by capsaicin was associated with anti-cancer effects. Therefore, TRPV1 provides a potential link between the process of inflammation, cancer and immunity, and offers new treatment possibilities. Nevertheless, in many cases, results regarding TRPV1 are contradictory and need further refinement. In this review we present the summary of the data related to the role of TRPV1 channel in the process of inflammation, cancer and immunity, limitations of the studies, and directions for future research. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract We previously demonstrated that capsazepine (CPZ), a synthetic transient receptor potential Vanilloid subtype 1 (TRPV1) antagonist, has significant anti-cancer effects in vivo. The purpose of this study was to develop more potent analogs based upon CPZ pharmacophore and structure–activity relationships (SAR) across analogs. We generated 30 novel compounds and screened for their anti-proliferative effects in cultured HeLa cervical cancer cells. Cell viability assays identified multiple compounds with IC 50s < 15 μM and one compound, 29 with an IC 50 < 5 μM; six fold more potent than CPZ. We validated the anti-proliferative efficacy of two lead compounds, 17 and 29 , in vivo using HeLa-derived xenografts in athymic nude mice. Both analogs significantly reduced tumor volumes by day 8 compared to control treated animals (p < 0.001) with no observable adverse effects. Calcium imaging determined that compound 17 activates TRPV1 whereas 29 neither activates nor inhibits TRPV1; indicating a unique mechanism-of-action that does not involve TRPV1 signaling. Cell viability assays using a panel of additional tumor types including oral squamous cell carcinoma, non-small cell lung cancer (NSCLC), breast cancer, and prostate cancer cell lines (HSC-3, H460, MDA-231, and PC-3 respectively) demonstrated that both lead compounds were efficacious against every cancer type tested. Compounds 29 displayed IC 50s of 1–2.5 μM in HSC-3and PC-3cells. Thus, we propose that these novel CPZ analogs may serve as efficacious therapeutic agents against multiple tumor types that warrant further development for clinical application. [ABSTRACT FROM AUTHOR]

Capsazepine is a competitive antagonist of capsaicin, a TRPV1 agonist responsible for the spicy taste of pepper. TRPV1 agonists and antagonists are known to affect mammalian body temperature, but their action on thermoregulation in insects is poorly known. In this study we evaluated the effect of capsazepine on the thermal preference of the American cockroach, Periplaneta americana using a thermal gradient. Our results revealed that capsazepine in submicromolar concentrations induces a preference for higher ambient temperatures when compared to the control insects. To assess whether capsazepine may act also as an antagonist of capsaicin in insects, we determined this insects' thermal behaviour when capsazepine was applied before capsaicin. The hypothermic response to capsaicin was clearly blocked by pre-treatment with capsazepine only in female American cockroaches. Our results indicate the involvement of structures functionally similar to TRPV1 in insect thermosensation.

Intracellular calcium (Ca2+) concentration ([Ca2+]i) is a key determinant of cell fate and is implicated in carcinogenesis. Membrane ion channels are structures through which ions enter or exit the cell, depending on the driving forces. The opening of transient receptor potential vanilloid 1 (TRPV1) ligand-gated ion channels facilitates transmembrane Ca2+ and Na+ entry, which modifies the delicate balance between apoptotic and proliferative signaling pathways. Proliferation is upregulated through two mechanisms: (1) ATP binding to the G-protein-coupled receptor P2Y2, commencing a kinase signaling cascade that activates the serine-threonine kinase Akt, and (2) the transactivation of the epidermal growth factor receptor (EGFR), leading to a series of protein signals that activate the extracellular signal-regulated kinases (ERK) 1/2. The TRPV1-apoptosis pathway involves Ca2+ influx and efflux between the cytosol, mitochondria, and endoplasmic reticulum (ER), the release of apoptosis-inducing factor (AIF) and cytochrome c from the mitochondria, caspase activation, and DNA fragmentation and condensation. While proliferative mechanisms are typically upregulated in cancerous tissues, shifting the balance to favor apoptosis could support anti-cancer therapies. TRPV1, through [Ca2+]i signaling, influences cancer cell fate; therefore, the modulation of the TRPV1-enforced proliferation–apoptosis balance is a promising avenue in developing anti-cancer therapies and overcoming cancer drug resistance. As such, this review characterizes and evaluates the role of TRPV1 in cell death and survival, in the interest of identifying mechanistic targets for drug discovery.

Introduction Earlier reports on the effect of temperature on gut motility concentrated on experiments conducted on the small intestines of adult animals. The effect of temperature on the large intestine, particularly in neonates, warrants further investigation. The current study investigated the effect of a temperature increase and its mechanism in the colon and rectum of neonate and adult rats. Methods and materials In an organ bath preparation, segments from the colon and rectum were subjected to increasing bath temperatures (37°C-40°C). In other groups, pretreatment with capsazepine (1 µM) and Nω-nitro-l-arginine methyl ester (L-NAME) (100 µM) was done, in different groups, to assess their impact on temperature-induced contractile response. Results Increasing the bath temperature significantly reduced the contractile tension in the colon and rectum. When L-NAME (100 µM)-pretreated segments of the colon and rectum were subjected to different bath temperatures, the contractile tension increased compared to the contractile tension at different bath temperatures without any drug. Capsazepine (1 µM) pretreatment, on the other hand, enhanced the decrease in the contractile tension in the colon and rectum of adult rats compared to the contractile tension produced at different bath temperatures without any drug, while in neonates, capsazepine (1 µM) pretreatment caused a rise in the contractile tension in the rectum with no effect in the colon. Increased bath temperature from 37°C to 40°C increased the contractile frequency in the colon and rectum in both adult and neonate rats. Pretreatment with L-NAME (100 µM) and capsazepine (1 µM) in adults and L-NAME (100 µM) in neonates caused an increase in the contractile frequency in both the colon and rectum; on the other hand, capsazepine pretreatment did not affect the contractile frequency in the colon and rectum of neonate rats compared to the contractile frequency produced at different bath temperatures without any drug. Conclusion The contractile response of rats' large intestines, colon, and rectum to increasing temperatures may involve nitric oxide (NO)-mediated and transient receptor potential vanilloid-1 (TRPV1)-mediated mechanisms. The effects of capsazepine on the colon and rectum of adults and neonates differ, possibly due to the TRPV1-mediated mechanism not developing properly in the neonate and developing later in adulthood., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2023, Singh et al.)

Acute pancreatitis (AP) is the acute inflammation of the pancreas. The morbidity of AP has increased in recent years. Certain patients eventually develop severe AP (SAP), which rapidly progresses to multiple organ dysfunction; the incidence of this occurring in patients with AP is 20‑30%. To date, no specific drugs or methods exist to treat this disease. Rutaecarpine relaxes vascular smooth muscle by stimulating calcitonin gene‑related peptide (CGRP) release via activation of vanilloid receptor subtype 1 (VR1). It has been demonstrated that rutaecarpine induces a therapeutic effect on SAP. The present study was conducted to characterize the molecular mechanisms underlying the protective effects of rutaecarpine against AP using a rat model of AP. Gross pathological changes of the pancreas, as well as the pancreatic tissue histopathological score, were assessed following treatment with rutaecarpine, capsazepine or a combination of the two. Serum amylase activity was detected using an automatic biochemistry analyzer. Changes in the serum concentrations of interleukin (IL)‑6, tumor necrosis factor (TNF‑α), IL‑10 and CGRP were assessed by ELISA and radioimmunoassay. The results demonstrated that pre‑treatment with rutaecarpine markedly decreased pancreatic inflammation and necrosis, reduced the volume of ascites, and significantly increased the plasma concentration of CGRP and the serum concentration of IL‑10, an anti‑inflammatory cytokine. However, serum concentrations of the inflammatory cytokines IL‑6 and TNF‑α were decreased. The effect of rutaecarpine treatment markedly improved with increases in the drug dose. Capsazepine, as a competitive vanilloid receptor antagonist, abolished these protective effects of rutaecarpine against AP. Therefore, the results of the present study indicate that rutaecarpine protects against AP in rats by upregulating endogenous CGRP release via activation VR1 of, to improving the microcirculation of the pancreatic tissue and regulate the expression of inflammatory factors. [ABSTRACT FROM AUTHOR]

Aims Intracellular calcium plays an important role in neuronal hyperexcitability that leads to seizures. One calcium influx route of interest is the transient receptor potential vanilloid type 1 ( TRPV1) channel. Here, we evaluated the effects of capsazepine ( CPZ), a potent blocker of TRPV1 channels on acoustically evoked seizures (audiogenic seizures, AGS) in the genetically epilepsy-prone rat ( GEPR-3), a model of inherited epilepsy. Methods Male and female GEPR-3s were used. For the acute CPZ treatment study, GEPR-3s were tested for AGS susceptibility before and after treatment with various doses of CPZ (0, 1, 3, and 10 mg/kg; ip). For semichronic CPZ treatment study, GEPR-3s were tested for AGS susceptibility before and after 5-day CPZ treatment at the dose of 1 mg/kg (ip). The prevalence, latency, and severity of AGS were recorded and analyzed. Results We found that acute CPZ pretreatment reduced the seizure severity in male GEPR-3s; the effect was dose-dependent. In female GEPR-3s, however, CPZ treatment completely suppressed the seizure susceptibility. Furthermore, semichronic CPZ treatment suppressed seizure susceptibility in female GEPR-3s, but only reduced the seizure severity in male GEPR-3s. Conclusions These findings suggest that the TRPV1 channel is a promising molecular target for seizure suppression, with female GEPR-3s exhibiting higher sensitivity than male GEPR-3s. [ABSTRACT FROM AUTHOR]

Aim Ethanol is one of the most widely used recreational drugs in the world. At high concentrations, it can induce carotid artery vasoconstriction. Hyperthermia potentiates its effects resulting in carotid artery vasoconstriction at any concentration. The aim of this study is to investigate the interaction between ethanol and heating and to understand the underlying mechanisms leading to their synergistic effect. Materials and methods Isometric tension of rabbit carotid artery ring segments suspended in organ baths filled with Krebs solution was recorded. Different concentrations of ethanol were examined at 37 °C and during temperature elevation to39–43 °C. Capsaicin and capsazepine were used to examine the mechanism of action of ethanol. Key findings Ethanol induced contraction at 37 °C when the concentration reached 100 mM. Contraction was observed at any concentration at higher temperatures. Ethanol potentiated heat-induced contraction. Capsaicin, the vanilloid receptor subtype1 (TRPV1) agonist, potentiated the vasoconstriction due to heating. While capsazepine, TRPV1 antagonist, abolished the effect of ethanol and its potentiation of heating-induced contraction, but it did not abolish the heating effect. Significance Ethanol's mechanism of action and its effect on heating induced-vasoconstriction of the carotid artery is being mediated by TRPV1. The combination of ethanol and hyperthermia can lead to a synergistic effect on carotid vasoconstriction. This effect may induce brain damage and heat stroke. Development of new drugs act as TRPV1 antagonist can be used to prevent these fatal effects. [ABSTRACT FROM AUTHOR]

• The presence of TRPV1 channels has been demonstrated in many regions of the brain. • TRPV1 channels are involved in synaptic transmission and cognitive functions. • TRPV1 channels are associated with neuroinflammation and neurodegenerative process. • TRPV1 agonist capsaicin attenuates OKA-induced Alzheimer's Disease-like pathology. The presence of Transient Receptor Potential Vanilloid 1 (TRPV1) channels was detected in many regions of the human and rat brain, including the cortex and hippocampus. TRPV1 channels have functions such as the modulation of synaptic transmission and plasticity and the regulation of cognitive functions. Previous studies conducted with TRPV1 agonists and antagonists show that this channel is associated with the neurodegenerative process. In the present study, the purpose was to investigate the effects of capsaicin, which is a TRPV1 agonist, and capsazepine, a TRPV1 antagonist, in the Alzheimer's Disease (AD) model that was induced by intracerebroventricular (ICV) administration of okadaic acid (OKA). The AD-like experimental model was created with bilateral ICV OKA injection. Intraperitoneal capsaicin and capsazepine injections were administered to the treatment groups for 13 days and histological and immunohistochemical examinations were performed from the cortex and hippocampal CA3 regions of the brain. The Morris Water Maze Test was used for spatial memory measurement. ICV OKA administration increased the levels of caspase-3, phosphorylated-tau-(ser396), Aβ, TNF-α, and IL1-β, from the cortex and hippocampal CA3 regions of the brain and decreased the phosphorylated-Glycogen synthase kinase-3 beta-(ser9) levels. In addition, the OKA administration corrupted the spatial memory. The TRPV1 agonist capsaicin reversed the pathological changes induced by ICV OKA administration, but not the TRPV1 antagonist capsazepine. It was found in the study that the administration of the TRPV1 agonist capsaicin reduced neurodegeneration, neuroinflammation, and deterioration in spatial memory in the AD model induced by OKA. [ABSTRACT FROM AUTHOR]

Capsazepine is a synthetic analogue of capsaicin that can function as an antagonist of TRPV1. Capsazepine can exhibit diverse effects on cancer (prostate cancer, breast cancer, colorectal cancer, oral cancer, and osteosarcoma) growth and survival, and can be therapeutically used against other major disorders such as colitis, pancreatitis, malaria, and epilepsy. Capsazepine has been reported to exhibit pleiotropic anti-cancer effects against numerous tumor cell lines. Capsazepine can modulate Janus activated kinase (JAK)/signal transducer and activator of the transcription (STAT) pathway, intracellular Ca2+ concentration, and reactive oxygen species (ROS)-JNK-CCAAT/enhancer-binding protein homologous protein (CHOP) pathways. It can inhibit cell proliferation, metastasis, and induce apoptosis. Moreover, capsazepine can exert anti-inflammatory effects through the downregulation of lipopolysaccharide (LPS)-induced nuclear transcription factor-kappa B (NF-κB), as well as the blockage of activation of both transient receptor potential cation channel subfamily V member 1 (TRPV1) and transient receptor potential cation channel, subfamily A, and member 1 (TRPA1). This review briefly summarizes the diverse pharmacological actions of capsazepine against various cancers and inflammatory conditions.

Asthma is a common disorder characterized, in part, by airway smooth muscle (ASM) hyperresponsiveness. Transient receptor potential vanilloid 1 (TRPV1) is a nonselective cation channel expressed on airway nerve fibers that modulates afferent signals, resulting in cough, and potentially bronchoconstriction. In the present study, the TRPV1 transcript was detected by RT-PCR in primary cultured human ASM cells, and the TRPV1 protein was detected in ASM of human trachea by immunohistochemistry. Proximity ligation assays suggest that TRPV1 is expressed in the sarcoplasmic reticulum membrane of human ASM cells in close association with sarco/endoplasmic reticulum Ca2+-ATPase-2. In guinea pig tracheal ring organ bath experiments, the TRPV1 agonist capsaicin led to ASM contraction, but this contraction was significantly attenuated by the sodium channel inhibitor bupivacaine (n = 4, P < 0.05) and the neurokinin-2 receptor antagonist GR-159897 (n = 4, P < 0.05), suggesting that this contraction is neutrally mediated. However, pretreatment of guinea pig and human ASM in organ bath experiments with the TRPV1 antagonist capsazepine inhibited the maintenance phase of an acetylcholine-induced contraction (n = 4, P < 0.01 for both species). Similarly, capsazepine inhibited methacholine-induced contraction of peripheral airways in mouse precision-cut lung slice (PCLS) experiments (n = 4–5, P < 0.05). Although capsazepine did not inhibit store-operated calcium entry in mouse ASM cells in PCLS (n = 4–7, P = nonsignificant), it did inhibit calcium oscillations (n = 3, P < 0.001). These studies suggest that TRPV1 is expressed on ASM, including the SR, but that ASM TRPV1 activation does not play a significant role in initiation of ASM contraction. However, capsazepine does inhibit maintenance of contraction, likely by inhibiting calcium oscillations. [ABSTRACT FROM AUTHOR]

Transient receptor potential vanilloid member 1 (TRPV1) is a heat and capsaicin receptor that allows cations to permeate and cause pain. As the molecular basis for temperature sensing, the heat capacity (Δ C p , 19492-19497 (2011).] has been proposed and experimentally supported. Theoretically, heat capacity is proportional to a variance in enthalpy, presumably related to structural fluctuation; however, the fluctuation of TRPV1 has not been directly visualized. In this study, we directly visualized single-molecule structural fluctuations of the TRPV1 channels in a lipid bilayer with the ligands resiniferatoxin (agonist, 1,000 times hotter than capsaicin) and capsazepine (antagonist) by high-speed atomic force microscopy. We observed the structural fluctuations of TRPV1 in an apo state and found that RTX binding enhances structural fluctuations, while CPZ binding suppresses fluctuations. These ligand-dependent differences in structural fluctuation would play a key role in the gating of TRPV1.Proc. Natl. Acad. Sci. U.S.A. 108 , 19492-19497 (2011).] has been proposed and experimentally supported. Theoretically, heat capacity is proportional to a variance in enthalpy, presumably related to structural fluctuation; however, the fluctuation of TRPV1 has not been directly visualized. In this study, we directly visualized single-molecule structural fluctuations of the TRPV1 channels in a lipid bilayer with the ligands resiniferatoxin (agonist, 1,000 times hotter than capsaicin) and capsazepine (antagonist) by high-speed atomic force microscopy. We observed the structural fluctuations of TRPV1 in an apo state and found that RTX binding enhances structural fluctuations, while CPZ binding suppresses fluctuations. These ligand-dependent differences in structural fluctuation would play a key role in the gating of TRPV1.

Capsazepine is a competitive antagonist of capsaicin, a TRPV1 agonist responsible for the spicy taste of pepper. TRPV1 agonists and antagonists are known to affect mammalian body temperature, but their action on thermoregulation in insects is poorly known. In this study we evaluated the effect of capsazepine on the thermal preference of the American cockroach, Periplaneta americana using a thermal gradient. Our results revealed that capsazepine in submicromolar concentrations induces a preference for higher ambient temperatures when compared to the control insects. To assess whether capsazepine may act also as an antagonist of capsaicin in insects, we determined this insects' thermal behaviour when capsazepine was applied before capsaicin. The hypothermic response to capsaicin was clearly blocked by pre-treatment with capsazepine only in female American cockroaches. Our results indicate the involvement of structures functionally similar to TRPV1 in insect thermosensation. [ABSTRACT FROM AUTHOR]