WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells
The CB<sub>1</sub> cannabinoid receptor (CB<sub>1</sub>R) and extracellular calcium (eCa<sup>2+</sup>)-stimulated Calcium Sensing receptor (CaSR) can exert cellular signaling by modulating levels of intracellular calcium ([Ca<sup>2+</sup>]<sub>i&...
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2022-09-01
|
| Series: | Cells |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2073-4409/11/19/2947 |
| _version_ | 1797480092709945344 |
|---|---|
| author | Victor M. Pulgar Allyn C. Howlett Khalil Eldeeb |
| author_facet | Victor M. Pulgar Allyn C. Howlett Khalil Eldeeb |
| author_sort | Victor M. Pulgar |
| collection | DOAJ |
| description | The CB<sub>1</sub> cannabinoid receptor (CB<sub>1</sub>R) and extracellular calcium (eCa<sup>2+</sup>)-stimulated Calcium Sensing receptor (CaSR) can exert cellular signaling by modulating levels of intracellular calcium ([Ca<sup>2+</sup>]<sub>i</sub>). We investigated the mechanisms involved in the ([Ca<sup>2+</sup>]<sub>i</sub>) increase in N18TG2 neuroblastoma cells, which endogenously express both receptors. Changes in [Ca<sup>2+</sup>]<sub>i</sub> were measured in cells exposed to 0.25 or 2.5 mM eCa<sup>2+</sup> by a ratiometric method (Fura-2 fluorescence) and expressed as the difference between baseline and peak responses (ΔF<sub>340/380</sub>). The increased ([Ca<sup>2+</sup>]<sub>i</sub>) in cells exposed to 2.5 mM eCa<sup>2+</sup> was blocked by the CaSR antagonist, NPS2143, this inhibition was abrogated upon stimulation with WIN55212-2. WIN55212-2 increased [Ca<sup>2+</sup>]<sub>i</sub> at 0.25 and 2.5 mM eCa<sup>2+</sup> by 700% and 350%, respectively, but this increase was not replicated by CP55940 or methyl-anandamide. The store-operated calcium entry (SOCE) blocker, MRS1845, attenuated the WIN55212-2-stimulated increase in [Ca<sup>2+</sup>]<sub>i</sub> at both levels of eCa<sup>2+</sup>. Simultaneous perfusion with the CB<sub>1</sub> antagonist, SR141716 or NPS2143 decreased the response to WIN55212-2 at 0.25 mM but not 2.5 mM eCa<sup>2+</sup>. Co-perfusion with the non-CB<sub>1</sub>/CB<sub>2</sub> antagonist O-1918 attenuated the WIN55212-2-stimulated [Ca<sup>2+</sup>]<sub>i</sub> increase at both eCa<sup>2+</sup> levels. These results are consistent with WIN55212-2-mediated intracellular Ca<sup>2+</sup> mobilization from store-operated calcium channel-filled sources that could occur via either the CB<sub>1</sub>R or an O-1918-sensitive non-CB<sub>1</sub>R in coordination with the CaSR. Intracellular pathway crosstalk or signaling protein complexes may explain the observed effects. |
| first_indexed | 2024-03-09T21:55:10Z |
| format | Article |
| id | doaj.art-dd079d48601f46178c770fb42f96ba9b |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-09T21:55:10Z |
| publishDate | 2022-09-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-dd079d48601f46178c770fb42f96ba9b2023-11-23T19:59:59ZengMDPI AGCells2073-44092022-09-011119294710.3390/cells11192947WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma CellsVictor M. Pulgar0Allyn C. Howlett1Khalil Eldeeb2Department of Pharmaceutical and Clinical Sciences, College of Pharmacy and Health Sciences, Campbell University, Buies Creek, NC 27506, USADepartment of Physiology & Pharmacology, Wake Forest School of Medicine, Winston-Salem, NC 27157, USADepartment of Physiology & Pharmacology, Wake Forest School of Medicine, Winston-Salem, NC 27157, USAThe CB<sub>1</sub> cannabinoid receptor (CB<sub>1</sub>R) and extracellular calcium (eCa<sup>2+</sup>)-stimulated Calcium Sensing receptor (CaSR) can exert cellular signaling by modulating levels of intracellular calcium ([Ca<sup>2+</sup>]<sub>i</sub>). We investigated the mechanisms involved in the ([Ca<sup>2+</sup>]<sub>i</sub>) increase in N18TG2 neuroblastoma cells, which endogenously express both receptors. Changes in [Ca<sup>2+</sup>]<sub>i</sub> were measured in cells exposed to 0.25 or 2.5 mM eCa<sup>2+</sup> by a ratiometric method (Fura-2 fluorescence) and expressed as the difference between baseline and peak responses (ΔF<sub>340/380</sub>). The increased ([Ca<sup>2+</sup>]<sub>i</sub>) in cells exposed to 2.5 mM eCa<sup>2+</sup> was blocked by the CaSR antagonist, NPS2143, this inhibition was abrogated upon stimulation with WIN55212-2. WIN55212-2 increased [Ca<sup>2+</sup>]<sub>i</sub> at 0.25 and 2.5 mM eCa<sup>2+</sup> by 700% and 350%, respectively, but this increase was not replicated by CP55940 or methyl-anandamide. The store-operated calcium entry (SOCE) blocker, MRS1845, attenuated the WIN55212-2-stimulated increase in [Ca<sup>2+</sup>]<sub>i</sub> at both levels of eCa<sup>2+</sup>. Simultaneous perfusion with the CB<sub>1</sub> antagonist, SR141716 or NPS2143 decreased the response to WIN55212-2 at 0.25 mM but not 2.5 mM eCa<sup>2+</sup>. Co-perfusion with the non-CB<sub>1</sub>/CB<sub>2</sub> antagonist O-1918 attenuated the WIN55212-2-stimulated [Ca<sup>2+</sup>]<sub>i</sub> increase at both eCa<sup>2+</sup> levels. These results are consistent with WIN55212-2-mediated intracellular Ca<sup>2+</sup> mobilization from store-operated calcium channel-filled sources that could occur via either the CB<sub>1</sub>R or an O-1918-sensitive non-CB<sub>1</sub>R in coordination with the CaSR. Intracellular pathway crosstalk or signaling protein complexes may explain the observed effects.https://www.mdpi.com/2073-4409/11/19/2947Ca<sup>2+</sup> mobilizationreceptor crosstalkcannabimimetic aminoalkylindoles |
| spellingShingle | Victor M. Pulgar Allyn C. Howlett Khalil Eldeeb WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells Cells Ca<sup>2+</sup> mobilization receptor crosstalk cannabimimetic aminoalkylindoles |
| title | WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells |
| title_full | WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells |
| title_fullStr | WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells |
| title_full_unstemmed | WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells |
| title_short | WIN55212-2 Modulates Intracellular Calcium via CB<sub>1</sub> Receptor-Dependent and Independent Mechanisms in Neuroblastoma Cells |
| title_sort | win55212 2 modulates intracellular calcium via cb sub 1 sub receptor dependent and independent mechanisms in neuroblastoma cells |
| topic | Ca<sup>2+</sup> mobilization receptor crosstalk cannabimimetic aminoalkylindoles |
| url | https://www.mdpi.com/2073-4409/11/19/2947 |
| work_keys_str_mv | AT victormpulgar win552122modulatesintracellularcalciumviacbsub1subreceptordependentandindependentmechanismsinneuroblastomacells AT allynchowlett win552122modulatesintracellularcalciumviacbsub1subreceptordependentandindependentmechanismsinneuroblastomacells AT khalileldeeb win552122modulatesintracellularcalciumviacbsub1subreceptordependentandindependentmechanismsinneuroblastomacells |