Large conductance calcium-and voltage-gated potassium (BK) channels are important regulators of physiological homeostasis and their function is potently modulated by protein kinase A (PKA) phosphorylation. PKA regulates the channel through phosphorylation of residues within the intracellular C terminus of the poreforming ␣-subunits. However, the molecular mechanism(s) by which phosphorylation of the ␣-subunit effects changes in channel activity are unknown. Inhibition of BK channels by PKA depends on phosphorylation of only a single ␣-subunit in the channel tetramer containing an alternatively spliced insert (STREX) suggesting that phosphorylation results in major conformational rearrangements of the C terminus. Here, we define the mechanism of PKA inhibition of BK channels and demonstrate that this regulation is conditional on the palmitoylation status of the channel. We show that the cytosolic C terminus of the STREX BK channel uniquely interacts with the plasma membrane via palmitoylation of evolutionarily conserved cysteine residues in the STREX insert. PKA phosphorylation of the serine residue immediately upstream of the conserved palmitoylated cysteine residues within STREX dissociates the C terminus from the plasma membrane, inhibiting STREX channel activity. Abolition of STREX palmitoylation by site-directed mutagenesis or pharmacological inhibition of palmitoyl transferases prevents PKA-mediated inhibition of BK channels. Thus, palmitoylation gates BK channel regulation by PKA phosphorylation. Palmitoylation and phosphorylation are both dynamically regulated; thus, cross-talk between these 2 major posttranslational signaling cascades provides a mechanism for conditional regulation of BK channels. Interplay of these distinct signaling cascades has important implications for the dynamic regulation of BK channels and physiological homeostasis.L arge conductance calcium-and voltage-gated potassium (BK) channels are potently regulated by protein phosphorylation (1) and are important determinants of neuronal, cardiovascular, endocrine, and epithelial function where channel dysfunction may lead to major disorders such as hypertension (2, 3), ataxia (4), epilepsy (5, 6), and incontinence (7). BK channels are potently regulated by phosphorylation, and several putative phosphorylation motifs on the pore-forming ␣-subunit have been identified (8-12). However, as for other potassium channels, the molecular basis through which phosphorylation of the ␣-subunit effects changes in BK channel activity is essentially unknown.BK channel pore-forming ␣-subunits are encoded by a single gene, KCNMA1 (13), and native BK channels show functional heterogeneity in their response to protein kinase A (PKA)-mediated phosphorylation. This diversity results, in large part, from the extensive alternative pre-mRNA splicing of the pore-forming ␣-subunits (10, 12). Previous studies have demonstrated that PKA phosphorylation of a conserved C-terminal phosphorylation motif. RQPS 899 results in BK channel activation (9,10,14). Inclusion of ...

These results indicate that a non-thermal cooling mouth rinse lowered thermal sensation, resulting in an elevated work rate, which extended exercise time in the heat at a fixed RPE.

PurposeThis study investigated the effects of l-menthol mouth rinse and ice slurry ingestion on time to exhaustion, when administered at the latter stages (~ 85%) of baseline exercise duration in the heat (35 °C).MethodTen male participants performed four time to exhaustion (TTE) trials on a cycle ergometer at 70% Wmax. In a randomized crossover design, (1) placebo-flavored non-calorific mouth rinse, (2) l-menthol mouth rinse (0.01%), or (3) ice ingestion (1.25 g kg−1), was administered at 85% of participants’ baseline TTE. Time to exhaustion, core and skin temperature, heart rate, rating of perceived effort, thermal comfort and thermal sensation were recorded.ResultsFrom the point of administration at 85% of baseline TTE, exercise time was extended by 1% (placebo, 15 s), 6% (l-menthol, 82 s) and 7% (ice, 108 s), relative to baseline performance (P = 0.036), with no difference between l-menthol and ice (P > 0.05). Core temperature, skin temperature, and heart rate increased with time but did not differ between conditions (P > 0.05). Thermal sensation did not differ significantly but demonstrated a large effect size (P = 0.080;  = 0.260).ConclusionThese results indicate that both thermally cooling and non-thermally cooling oral stimuli have an equal and immediate behavioral, rather than physiological, influence on exhaustive exercise in the heat.

Palmitoylation is emerging as an important and dynamic regulator of ion channel function; however, the specificity with which the large family of acyl palmitoyltransferases (zinc finger Asp-His-His-Cys type-containing acyl palmitoyltransferase (DHHCs)) control channel palmitoylation is poorly understood. We have previously demonstrated that the alternatively spliced stress-regulated exon (STREX) variant of the intracellular C-terminal domain of the large conductance calcium- and voltage-activated potassium (BK) channels is palmitoylated and targets the STREX domain to the plasma membrane. Using a combined imaging, biochemical, and functional approach coupled with loss-of-function (small interfering RNA knockdown of endogenous DHHCs) and gain-of-function (overexpression of recombinant DHHCs) assays, we demonstrate that multiple DHHCs control palmitoylation of the C terminus of STREX channels, the association of the STREX domain with the plasma membrane, and functional channel regulation. Cysteine residues 12 and 13 within the STREX insert were the only endogenously palmitoylated residues in the entire C terminus of the STREX channel. Palmitoylation of this dicysteine motif was controlled by DHHCs 3, 5, 7, 9, and 17, although DHHC17 showed the greatest specificity for this site upon overexpression of the cognate DHHC. DHHCs that palmitoylated the channel also co-assembled with the channel in co-immunoprecipitation experiments, and knockdown of any of these DHHCs blocked regulation of the channel by protein kinase A-dependent phosphorylation. Taken together our data reveal that a subset of DHHCs controls STREX palmitoylation and function and suggest that DHHC17 may preferentially target cysteine-rich domains. Finally, our approach may prove useful in elucidating the specificity of DHHC palmitoylation of intracellular domains of other ion channels and transmembrane proteins.

Objectives: Menthol is an organic compound with non-thermal cooling properties that has been shown to relieve thermal strain associated with exercise in the heat; however, its effects on performance have not been systematically analysed. The aims were to determine the effects of menthol applied (1) internally and (2) externally on exercise performance and thermal sensation. Design: Meta-analysisMethods: A search was performed using various databases in August 2018. The studies were screened using search criteria for eligibility. Thirteen peer-reviewed articles were identified for inclusion in a primary analysis on the effect of menthol on exercise performance; subsequently eleven of these articles were included in a secondary analysis on the effect of menthol on thermal sensation during exercise. A sub-analysis examining the application method was also performed.Results: Menthol improved overall exercise performance (Hedges' g = 0.33, 95 % CI -0.00, 0.65 P = 0.05), demonstrating greater effects when applied internally (Hedges' g = 0.40, 95 % CI 0.04, 0.76, P = 0.03). Thermal sensation was also lowered overall across all studies (Hedges' g = -0.54, 95 % CI -0.67, -0.42, P < 0.001). Conclusions:Exercise performance can be improved by application of non-thermally cooling menthol, which also reduces perceptual measures of thermal sensation.