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Sex Steroids Do Not Modulate TRPM2-Mediated Injury in Females following Middle Cerebral Artery Occlusion(1,2,3).

Quillinan N, Grewal H, Klawitter J, Herson PS - eNeuro (2014)

Bottom Line: Interestingly, treatment of female mice with the potent androgen receptor agonist DHT had no effect on ischemic injury and did not permit CTZ neuroprotection.Similarly, DHT-treated females did not exhibit increased levels of ADPribose, the TRPM2 ligand generated by PARP, following ischemia.In conclusion, our data demonstrate that while circulating androgens are necessary for PARP-mediated TRPM2 injury in males, they are not sufficient to produce TRPM2 activation in females.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, University of Colorado School of Medicine, Aurora, Colorado 80045.

ABSTRACT
Calcium-permeable transient receptor potential M2 (TRPM2) ion channel activation contributes to cerebral ischemic injury specifically in males. In male mice, circulating androgens are required for TRPM2 inhibition with clotrimazole (CTZ) to provide protection following experimental stroke. Sufficient levels of circulating androgens are necessary to support ischemia-induced activation of poly ADP ribose polymerase (PARP) and consequent activation of TRPM2 channels. In this study, we tested whether differences in sex steroids contribute to the lack of CTZ neuroprotection in females. Middle cerebral artery occlusion (MCAO) was performed using adult female mice that were hormonally intact, ovariectomized (OVX) or dihydrotestosterone (DHT) treated. CTZ or vehicle was administered at the time of reperfusion, animals were euthanized 24 h later and brains and serum were collected. Infarct analysis revealed no effect of CTZ in intact females or females lacking endogenous sex steroids (OVX). Interestingly, treatment of female mice with the potent androgen receptor agonist DHT had no effect on ischemic injury and did not permit CTZ neuroprotection. Similarly, DHT-treated females did not exhibit increased levels of ADPribose, the TRPM2 ligand generated by PARP, following ischemia. No differences in TRPM2 or androgen receptor expression were observed between males and females. These data suggest that the lack of TRPM2 activation in females following experimental stroke is not due to the presence of estrogen or the absence of androgens. In conclusion, our data demonstrate that while circulating androgens are necessary for PARP-mediated TRPM2 injury in males, they are not sufficient to produce TRPM2 activation in females.

No MeSH data available.


Related in: MedlinePlus

Clotrimazole is neuroprotective specifically in males. A, TTC staining and infarct analysis was performed 24 h after MCAO and presented as the percentage of corrected hemisphere. Clotrimazole administered at the time of reperfusion caused a significant reduction in infarct size in males but not females. Bars represent mean ± SEM. Comparisons made using one-way ANOVA and Tukey’s post hoc analysis; *p = 0.02 B, TRPM2 mRNA expression was measured by qRT-PCR and quantified relative to the housekeeping gene 18s. Data were normalized to male TRPM2 expression levels. No significant differences in TRPM2 mRNA were observed between males and females (Student’s t test; p = 0.12).
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Figure 1: Clotrimazole is neuroprotective specifically in males. A, TTC staining and infarct analysis was performed 24 h after MCAO and presented as the percentage of corrected hemisphere. Clotrimazole administered at the time of reperfusion caused a significant reduction in infarct size in males but not females. Bars represent mean ± SEM. Comparisons made using one-way ANOVA and Tukey’s post hoc analysis; *p = 0.02 B, TRPM2 mRNA expression was measured by qRT-PCR and quantified relative to the housekeeping gene 18s. Data were normalized to male TRPM2 expression levels. No significant differences in TRPM2 mRNA were observed between males and females (Student’s t test; p = 0.12).

Mentions: MCAO (60 min) was performed on adult male and female mice. The TRPM2 inhibitor clotrimazole (CTZ; 30 mg/kg, s.c.) or vehicle were administered at the time of reperfusion. Mice were euthanized 24 h after MCAO and TTC staining was performed on 2 mm brain sections (Fig. 1A). Infarct volume was smaller in CTZ-treated males (26.11 ± 2.25% of hemisphere, n = 8) compared to vehicle-treated males (38.98 ± 3.7%; n = 8; p = 0.02)a (Fig. 1B). In contrast, CTZ had no effect on infarct volume in intact female mice (29.85 ± 2.04%, n = 12 and 30.24 ± 2.83, n = 11, p = 0.99, respectively)b. Infarct volumes in vehicle-treated male and females were not significantly different (male vehicle vs female vehicle: p = 0.09c; male CTZ vs female CTZ: p = 0.76d). The lack of protection observed in CTZ-treated female mice is not the result of reduced expression in females as mRNA levels of TRPM2 transcript were not different from males (1.000 ± 0.07692, n = 4) and females (1.274 ± 0.1219, n = 5; p = 0.11e).


Sex Steroids Do Not Modulate TRPM2-Mediated Injury in Females following Middle Cerebral Artery Occlusion(1,2,3).

Quillinan N, Grewal H, Klawitter J, Herson PS - eNeuro (2014)

Clotrimazole is neuroprotective specifically in males. A, TTC staining and infarct analysis was performed 24 h after MCAO and presented as the percentage of corrected hemisphere. Clotrimazole administered at the time of reperfusion caused a significant reduction in infarct size in males but not females. Bars represent mean ± SEM. Comparisons made using one-way ANOVA and Tukey’s post hoc analysis; *p = 0.02 B, TRPM2 mRNA expression was measured by qRT-PCR and quantified relative to the housekeeping gene 18s. Data were normalized to male TRPM2 expression levels. No significant differences in TRPM2 mRNA were observed between males and females (Student’s t test; p = 0.12).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4596140&req=5

Figure 1: Clotrimazole is neuroprotective specifically in males. A, TTC staining and infarct analysis was performed 24 h after MCAO and presented as the percentage of corrected hemisphere. Clotrimazole administered at the time of reperfusion caused a significant reduction in infarct size in males but not females. Bars represent mean ± SEM. Comparisons made using one-way ANOVA and Tukey’s post hoc analysis; *p = 0.02 B, TRPM2 mRNA expression was measured by qRT-PCR and quantified relative to the housekeeping gene 18s. Data were normalized to male TRPM2 expression levels. No significant differences in TRPM2 mRNA were observed between males and females (Student’s t test; p = 0.12).
Mentions: MCAO (60 min) was performed on adult male and female mice. The TRPM2 inhibitor clotrimazole (CTZ; 30 mg/kg, s.c.) or vehicle were administered at the time of reperfusion. Mice were euthanized 24 h after MCAO and TTC staining was performed on 2 mm brain sections (Fig. 1A). Infarct volume was smaller in CTZ-treated males (26.11 ± 2.25% of hemisphere, n = 8) compared to vehicle-treated males (38.98 ± 3.7%; n = 8; p = 0.02)a (Fig. 1B). In contrast, CTZ had no effect on infarct volume in intact female mice (29.85 ± 2.04%, n = 12 and 30.24 ± 2.83, n = 11, p = 0.99, respectively)b. Infarct volumes in vehicle-treated male and females were not significantly different (male vehicle vs female vehicle: p = 0.09c; male CTZ vs female CTZ: p = 0.76d). The lack of protection observed in CTZ-treated female mice is not the result of reduced expression in females as mRNA levels of TRPM2 transcript were not different from males (1.000 ± 0.07692, n = 4) and females (1.274 ± 0.1219, n = 5; p = 0.11e).

Bottom Line: Interestingly, treatment of female mice with the potent androgen receptor agonist DHT had no effect on ischemic injury and did not permit CTZ neuroprotection.Similarly, DHT-treated females did not exhibit increased levels of ADPribose, the TRPM2 ligand generated by PARP, following ischemia.In conclusion, our data demonstrate that while circulating androgens are necessary for PARP-mediated TRPM2 injury in males, they are not sufficient to produce TRPM2 activation in females.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, University of Colorado School of Medicine, Aurora, Colorado 80045.

ABSTRACT
Calcium-permeable transient receptor potential M2 (TRPM2) ion channel activation contributes to cerebral ischemic injury specifically in males. In male mice, circulating androgens are required for TRPM2 inhibition with clotrimazole (CTZ) to provide protection following experimental stroke. Sufficient levels of circulating androgens are necessary to support ischemia-induced activation of poly ADP ribose polymerase (PARP) and consequent activation of TRPM2 channels. In this study, we tested whether differences in sex steroids contribute to the lack of CTZ neuroprotection in females. Middle cerebral artery occlusion (MCAO) was performed using adult female mice that were hormonally intact, ovariectomized (OVX) or dihydrotestosterone (DHT) treated. CTZ or vehicle was administered at the time of reperfusion, animals were euthanized 24 h later and brains and serum were collected. Infarct analysis revealed no effect of CTZ in intact females or females lacking endogenous sex steroids (OVX). Interestingly, treatment of female mice with the potent androgen receptor agonist DHT had no effect on ischemic injury and did not permit CTZ neuroprotection. Similarly, DHT-treated females did not exhibit increased levels of ADPribose, the TRPM2 ligand generated by PARP, following ischemia. No differences in TRPM2 or androgen receptor expression were observed between males and females. These data suggest that the lack of TRPM2 activation in females following experimental stroke is not due to the presence of estrogen or the absence of androgens. In conclusion, our data demonstrate that while circulating androgens are necessary for PARP-mediated TRPM2 injury in males, they are not sufficient to produce TRPM2 activation in females.

No MeSH data available.


Related in: MedlinePlus