J Vet Sci.  2016 Jun;17(2):137-144. 10.4142/jvs.2016.17.2.137.

Time-course changes of hippocalcin expression in the mouse hippocampus following pilocarpine-induced status epilepticus

Affiliations
  • 1Department of Pharmacy, College of Pharmacy, Dankook University, Cheonan 31116, Korea. anaphy@dankook.ac.kr

Abstract

Hippocalcin participates in the maintenance of neuronal calcium homeostasis. In the present study, we examined the time-course changes of neuronal degeneration and hippocalcin protein level in the mouse hippocampus following pilocarpine-induced status epilepticus (SE). Marked neuronal degeneration was observed in the hippocampus after SE in a time-dependent manner, although neuronal degeneration differed according to the hippocampal subregions. Almost no hippocalcin immunoreactivity was detected in the pyramidal neurons of the cornu ammonis 1 (CA1) region from 6 h after SE. However, many pyramidal neurons in the CA2 region showed hippocalcin immunoreactivity until 24 h after SE. In the CA3 region, only a few hippocalcin immunoreactive cells were observed at 12 h after SE, and almost no hippocalcin immunoreactivity was observed in the pyramidal neurons from 24 h after SE. Hippocalcin immunoreactivity in the polymorphic cells of the dentate gyrus was markedly decreased from 6 h after SE. In addition, hippocalcin protein level in the hippocampus began to decrease from 6 h after SE, and was significantly decreased at 24 h and 48 h after pilocarpine-induced SE. These results indicate that marked reduction of hippocalcin level may be closely related to neuronal degeneration in the hippocampus following pilocarpine-induced SE.

Keyword

hippocalcin; hippocampus; neuronal degeneration; status epilepticus

MeSH Terms

Animals
*Gene Expression Regulation/drug effects
Hippocalcin/*genetics/metabolism
Hippocampus/*metabolism
Male
Mice
Mice, Inbred ICR
Nerve Degeneration/chemically induced/*physiopathology
Pilocarpine/pharmacology
Status Epilepticus/chemically induced/*physiopathology
Time Factors
Pilocarpine
Hippocalcin

Figure

  • Fig. 1 Fluoro-Jade B (F-J B) histofluorescence staining in the mouse hippocampus of the control- (A) and pilocarpine-treated (B–E) groups. Many F-J B positive cells were detected in the CA1 region from 12 h after SE (arrows), and F-J B positive cells markedly increased in most of the pyramidal neurons in the CA3 region 24 h and 48 h after status epilepticus (SE; asterisks). In the DG, F-J B positive cells began to be observed in the polymorphic cells from 6 h after SE (arrowheads). Scale Bar = 1,000 µm.

  • Fig. 2 Immunohistochemistry for hippocalcin in the mouse hippocampus of the control- (A) and pilocarpine-treated (B–E) groups. In the control-group, hippocalcin immunoreactivity is primarily detected in the pyramidal neurons of the CA1-3 regions. Hippocalcin immunoreactivity in the mouse hippocampus gradually decreased following pilocarpine-induced SE in a time-dependent manner. Scale Bar = 1,000 µm.

  • Fig. 3 Immunohistochemistry for hippocalcin in the CA1 region of the control- (A) and pilocarpine-treated (B–E) groups. In the control-group, intense hippocalcin immunoreactivity was observed in the pyramidal neurons of the stratum pyramidale (SP; asterisks). However, hippocalcin immunoreactivity in the SP was not detected from 6 h following pilocarpine-induced SE. SO, stratum oriens; SR, stratum radiatum. Scale Bar = 100 µm.

  • Fig. 4 Immunohistochemistry for hippocalcin in the CA2/3 region of the control- (A) and pilocarpine-treated (B–E) groups. Hippocalcin immunoreactivity in the stratum pyramidale (SP) of the CA3 region decreased markedly from 6 h following pilocarpine-induced SE (asterisks). However, many pyramidal neurons in the CA2 region showed hippocalcin immunoreactivity until 24 h after SE (arrows). At 48 h after SE, no hippocalcin immunoreactive cells were observed in the CA2/3 region. Scale Bar = 200 µm.

  • Fig. 5 Immunohistochemistry for hippocalcin in the DG of the control- (A) and pilocarpine-treated (B–E) groups. Hippocalcin immunoreactivity is easily detected in the polymorphic cells of the polymorphic layer (PL) of the control-group (arrows). Hippocalcin immunoreactivity in the polymorphic cells was markedly decreased at 6 h after pilocarpine-induced SE, and no hippocalcin immunoreactive cells were observed in the PL from 12 h after SE. ML, molecular layer; GCL, granule cell layer. Scale Bar = 200 µm.

  • Fig. 6 Western blot analysis of hippocalcin (23 kDa) in the mouse hippocampus derived from the control- and pilocarpine-treated groups. Relative optical density (ROD) as % values of the immunoblot band are also shown (*p < 0.05, significantly different from the control group). The bars indicate the means ± SEM.


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