Myrcene, Terpenes, and TrpV1

This study came out of the laboratory of experimental medicine in Honolulu Hawaii. As a general background it might be worth while to review Ca²⁺-calmodulin inhibition of this Ca²⁺ channel and and Trp family overview.

This group selected myrcene and and a mixture of strain inspired terpenes. The latter was identified as dominant terpenes in strains of cannabis used for analgesia.

Terpene mixture: α-bisabolol, α-humulene, α-pinene, β-Caryophyllene, β-myrcene, (+)-β-pinene, camphene, limonene, linalool, nerolidol.

Cannabinoid mixture: Cannabidivarin (CBDV),Cannacebichromene (CBC), Cannabidiol (CBD), Cannabidiolic Acid (CBDA), Cannabigerol (CBG), Cannabigerolic Acid (CBGA), Cannabinol (CBN).

From reference [1]

• (a) This image of fluor-4 responding to intracellular Ca2+ is one chosen from an Internet search as an example. When capsacin is added relative fluorescent units (RFU) increase.
• (b) When a synthetic mixture of terpenes and cannabinoids is added, RFU increases.
• (c) The cannabinoid mixture does not do much by itself.
• (d) The terpene mixture causes a quick and robust increase in intracellular Ca²⁺.

from reference [1]

• (c) Myrcene alone causes a little dip followed by a rapid rise in intracellular Ca²⁺ in both the wild type and genetically modified to over produce TrpV1 HEK cells. This is highly suggestive that Ca²⁺ is increasing by means other than TrpV1. [1]
• (d). The same phenomenon is seen with a lower concentration of myrcene. [1]
• (e-i) These panels showed other myrcene concentrations and some experiments in which capsaicin but not the saline solution used to dissolve the capsaicin decreased intrcellular Ca²⁺.
• (j-i) When there is no extracellular Ca²⁺, myrcene still increases intracellular Ca²⁺. There has been a report of the ryanodine receptor in renal epithelial cells from the rabbit.

Some, but not all, of the terpenes tested caused some change in intracellular Ca²⁺ in HEK cells. [1]

Another group looked at the affect of components of Monarda fistulosa essential oil. {2] Myrcene and other monoterpene components did not affect fluo-4 fluorescence in HEK cells over expressing TrpV1 and TrpV4 that exceeded that found in non transfected cells. [2] β-myrcene activates TRPA1 in this study. Relative changes in intracellular Ca2+ were 0.111 ± 0.003 and 0.005 ± 0.001 in TrpA1 expressing and non transfected cells. [2]. The TrpA`1 ligand carvacol elicited 0.340 ± 0.012 0.061 ± 0.018 in TrpA1 expressing and non transfected cells. [2]. The

Let’s take a pause!

The Ghosh group [2] claims that myrcene and other terpenes found in cannabis do not regulate TrpV1. The Jansen group [2] claims that they do. We have to remember that kidney epithelial cells absorb calcium from the lumen of the proximal tubules. These calcium absorbing proteins are regulated by hormones that may be in different amounts from one lot of serum to the next. We have parathyroid and 1,25-hydroxy vitamin D controlling Ca²⁺ reabsorption. [3]

Adapted from reference [3] CaSR, calcium sensing resceptor, NKCC, sodium, potassium, chloride cotransporter, ROM renal outer medulalary potassium channel, NCC, codium chloride cotransporter.

The phenotypes of the HEK cells used in these experiments was not that well described. We do not know if beta myrcene is interacting with other Ca2+ transporting systems. The simplist explanation is that Ca2+ increases independent of TrpV1 are toxicology artifacts. In a rat study 1,000 mg/kg caused nephrosis in the outermost OSOM was much more pronounced, with prominent S3 tubule dilation. [4] Mycene has a molecular weight of 136.238 g·mol⁻¹ If we assume that 1kg of rat has a fluid volume of 0.5 to 1 liter, we have a concentration o f 7 to 15 μM. The Jansen group [1]might have been looking at nothing more than a toxicity induced Ca²⁺ leak.

The Jansen group also used a technique called whole cell patch clamping and offered more convincing evidence that TrpV1 is gated by myrcene. The also performed some molecular modeling to predict how this interaction might occur. [1]

References

1. Jansen, C., Shimoda, L., Kawakami, J. K., Ang, L., Bacani, A. J., Baker, J. D., Badowski, C., Speck, M., Stokes, A. J., Small-Howard, A. L., & Turner, H. (2019). Myrcene and terpene regulation of TRPV1. Channels (Austin, Tex.), 13(1), PMC free article
2. Ghosh, M., Schepetkin, I. A., Özek, G., Özek, T., Khlebnikov, A. I., Damron, D. S., & Quinn, M. T. (2020). Essential Oils from Monarda fistulosa: Chemical Composition and Activation of Transient Receptor Potential A1 (TRPA1) Channels. Molecules (Basel, Switzerland), 25(21), 4873. PMC free article
3. Patel AM, Goldfarb S. Got calcium? Welcome to the calcium-alkali syndrome. J Am Soc Nephrol. 2010 Sep;21(9):1440-3. PMC free article
4. Cesta MF, Hard GC, Boyce JT, Ryan MJ, Chan PC, Sills RC. Complex histopathologic response in rat kidney to oral β-myrcene: an unusual dose-related nephrosis and low-dose alpha2u-globulin nephropathy. Toxicol Pathol. 2013;41(8):1068-77. PMC free article

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This study came out of the laboratory of experimental medicine in Honolulu Hawaii. As a general background it might be worth while to review Ca²⁺-calmodulin inhibition of this Ca2+ channel and and Trp family overview.

This group selected myrcene and and a mixture of strain inspired terpenes. The latter was identified as dominant terpenes in strains of cannabis used for analgesia.

Terpene mixture: α-bisabolol, α-humulene, α-pinene, β-Caryophyllene, β-myrcene, (+)-β-pinene, camphene, limonene, linalool, nerolidol.

Cannabinoid mixture: Cannabidivarin (CBDV),Cannacebichromene (CBC), Cannabidiol (CBD), Cannabidiolic Acid (CBDA), Cannabigerol (CBG), Cannabigerolic Acid (CBGA), Cannabinol (CBN).

From reference [1]

• (a) This image of fluor-4 responding to intracellular Ca2+ is one chosen from an Internet search as an example. When capsacin is added relative fluorescent units (RFU) increase.
• (b) When a synthetic mixture of terpenes and cannabinoids is added, RFU increases.
• (c) The cannabinoid mixture does not do much by itself.
• (d) The terpene mixture causes a quick and robust increase in intracellular Ca²⁺.

from reference [1]

• (c) Myrcene alone causes a little dip followed by a rapid rise in intracellular Ca²⁺ in both the wild type and genetically modified to over produce TrpV1 HEK cells. This is highly suggestive that Ca²⁺ is increasing by means other than TrpV1. [1]
• (d). The same phenomenon is seen with a lower concentration of myrcene. [1]
• (e-i) These panels showed other myrcene concentrations and some experiments in which capsaicin but not the saline solution used to disolve the capsaicin decreased intrcellular Ca²⁺.
• (j-i) When there is no extracellular Ca²⁺, myrcene still increases intracellular Ca²⁺. There has been a report of the ryanodine receptor in renal epithelial cells from the rabbit.

Some, but not all, of the terpenes tested caused some change in intracellular Ca²⁺ in HEK cells. [1]

Another group looked at the affect of components of Monarda fistulosa essential oil. {2] Myrcene and other monoterpene components did not affect fluo-4 fluorescence in HEK cells over expressing TrpV1 and TrpV4 that exceeded that found in non transfected cells. [2] β-myrcene activates TRPA1 in this study. Relative changes in intracellular Ca2+ were 0.111 ± 0.003 and 0.005 ± 0.001 in TrpA1 expressing and non transfected cells. [2]. The TrpA`1 ligand carvacol elicited 0.340 ± 0.012 0.061 ± 0.018 in TrpA1 expressing and non transfected cells. [2]. The

Let’s take a pause!

The Ghosh group [2] claims that myrcene and other terpenes found in cannabis do not regulate TrpV1. The Jansen group [2] claims that they do. We have to remember that kidney epithelial cells absorb calcium from the lumen of the proximal tubules. These calcium absorbing proteins are regulated by hormones that may be in different amounts from one lot of serum to the next. We have parathyroid and 1,25-hydroxy vitamin D controlling Ca²⁺ reabsorption. [3]

Adapted from reference [3] CaSR, calcium sensing resceptor, NKCC, sodium, potassium, chloride cotransporter, ROM renal outer medulalary potassium channel, NCC, codium chloride cotransporter.

The phenotypes of the HEK cells used in these experiments was not that well described. We do not know if

The Jansen group also used a technique called whole cell patch clamping and offered more convincing evidence that TrpV1 is gated by myrcene. The also performed some molecular modeling to predict how this interaction might occur. [1]

References

1. Jansen, C., Shimoda, L., Kawakami, J. K., Ang, L., Bacani, A. J., Baker, J. D., Badowski, C., Speck, M., Stokes, A. J., Small-Howard, A. L., & Turner, H. (2019). Myrcene and terpene regulation of TRPV1. Channels (Austin, Tex.), 13(1), PMC free article
2. Ghosh, M., Schepetkin, I. A., Özek, G., Özek, T., Khlebnikov, A. I., Damron, D. S., & Quinn, M. T. (2020). Essential Oils from Monarda fistulosa: Chemical Composition and Activation of Transient Receptor Potential A1 (TRPA1) Channels. Molecules (Basel, Switzerland), 25(21), 4873. PMC free article
3. Patel AM, Goldfarb S. Got calcium? Welcome to the calcium-alkali syndrome. J Am Soc Nephrol. 2010 Sep;21(9):1440-3. PMC free article
4. Cesta MF, Hard GC, Boyce JT, Ryan MJ, Chan PC, Sills RC. Complex histopathologic response in rat kidney to oral β-myrcene: an unusual dose-related nephrosis and low-dose alpha2u-globulin nephropathy. Toxicol Pathol. 2013;41(8):1068-77. PMC free article