Potentiation of nerve growth factor-induced neurite outgrowth in PC12 cells by ifenprodil: the role of sigma-1 and IP3 receptors - PubMed (original) (raw)

Potentiation of nerve growth factor-induced neurite outgrowth in PC12 cells by ifenprodil: the role of sigma-1 and IP3 receptors

Tamaki Ishima et al. PLoS One. 2012.

Abstract

In addition to both the α1 adrenergic receptor and N-methyl-D-aspartate (NMDA) receptor antagonists, ifenprodil binds to the sigma receptor subtypes 1 and 2. In this study, we examined the effects of ifenprodil on nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. Ifenprodil significantly potentiated NGF-induced neurite outgrowth, in a concentration-dependent manner. In contrast, the α1 adrenergic receptor antagonist, prazosin and the NMDA receptor NR2B antagonist, Ro 25-6981 did not alter NGF-induced neurite outgrowth. Potentiation of NGF-induced neurite outgrowth mediated by ifenprodil was significantly antagonized by co-administration of the selective sigma-1 receptor antagonist, NE-100, but not the sigma-2 receptor antagonist, SM-21. Similarly, ifenprodil enhanced NGF-induced neurite outgrowth was again significantly reduced by the inositol 1,4,5-triphosphate (IP(3)) receptor antagonists, xestospongin C and 2-aminoethoxydiphenyl borate (2-APB) treatment. Furthermore, BAPTA-AM, a chelator of intracellular Ca(2+), blocked the effects of ifenprodil on NGF-induced neurite outgrowth, indicating the role of intracellular Ca(2+) in the neurite outgrowth. These findings suggest that activation at sigma-1 receptors and subsequent interaction with IP(3) receptors may mediate the pharmacological effects of ifenprodil on neurite outgrowth.

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Conflict of interest statement

Competing Interests: Dr. Hashimoto is a member of the Editorial Board of PLoS ONE. This does not alter the authors' adherence to all the PLoS ONE policies on sharing data and materials. The other author has declared that no competing interests exist.

Figures

Figure 1

Figure 1. Effects of ifenprodil with or without sigma receptor antagonists on NGF-induced neurite outgrowth in PC12 cells.

In the presence of NGF (2.5 ng/ml), ifenprodil (0.1, 1.0, or 10 µM) with or without NE-100 (1.0 µM; a sigma-1 receptor antagonist), or SM-21 (10 µM; a sigma-2 receptor antagonist) were incubated with PC12 cells. Four days after incubation with test drugs, morphometric analysis was performed. The data show the mean ± SEM (n = 6–18). **P<0.01, ***p<0.001 when compared to the control group. ###p<0.001 when compared to the ifenprodil (10 µM)-treated group.

Figure 2

Figure 2. Representative photographs of MAP-2 and GAP-43 immunocytochemistry in PC12 cells.

In the presence of NGF (2.5 ng/ml), vehicle control or ifenprodil (10 µM) were incubated with PC12 cells. Four days after incubation with test drugs, MAP-2 and GAP-43 immunocytochemistry was performed. Arrowhead shows the cells with neurite outgrowth.

Figure 3

Figure 3. Effects of α1 adrenergic receptor antagonist and NR2B antagonist on NGF-induced neurite outgrowth in PC12 cells.

In the presence of NGF (2.5 ng/ml), vehicle, prazosin (10 µM), or Ro 25-6981 (10 µM) were incubated with PC12 cells. Four days after incubation with test drugs, morphometric analysis was performed. The data show the mean ± SEM (n = 6). NS: Not significance.

Figure 4

Figure 4. Effects of IP3 receptor antagonists on NGF-induced neurite outgrowth in PC12 cells.

(A): In the presence of NGF (2.5 ng/ml), vehicle, ifenprodil (10 µM), ifenprodil (10 µM)+xestospongin C (1.0 µM), xestospongin C (1.0 µM) were incubated with PC12 cells. (B): In the presence of NGF (2.5 ng/ml), vehicle, ifenprodil (10 µM), ifenprodil (10 µM)+2-APB (100 µM), or 2-APB (100 µM) were incubated in PC12 cells. Four days after incubation with test drugs, morphometric analysis was performed. The data show the mean ± SEM (n = 6). ***p<0.001 when compared with the ifenprodil (10 µM)-treated group.

Figure 5

Figure 5. Effects of BAPTA-AM on potentiation of NGF-induced neurite outgrowth by ifenprodil.

In the presence of NGF (2.5 ng/ml), vehicle, ifenprodil (10 µM), ifenprodil (10 µM)+BAPTA-AM (5.0 µM), or BAPTA-AM (5.0 µM) were incubated with PC12 cells. Four days after incubation with test drugs, morphometric analysis was performed. The data show the mean ± SEM (n = 6). ***p<0.001 when compared to the control group. ###p<0.001 when compared to the ifenprodil (10 µM)-treated group.

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