Limits...
What Elements of the Inflammatory System Are Necessary for Epileptogenesis In Vitro?(1,2).

Park KI, Dzhala V, Saponjian Y, Staley KJ - eNeuro (2015)

Bottom Line: Organotypic hippocampal brain slices can be maintained in culture independently of the systemic inflammatory system, and the rapid course of epileptogenesis in these cultures supports the idea that inflammation is not necessary for epilepsy.However, this preparation still retains key cellular inflammatory mediators.These data support the idea that although the inflammatory system, neurons, and glia share key intercellular signaling molecules, neither systemic nor CNS-specific cellular elements of the immune and inflammatory systems are necessary components of epileptogenesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurology, Massachusetts General Hospital , Boston, Massachusetts 02129 ; Seoul Paik Hospital, Inje University , Seoul 100-032, South Korea.

ABSTRACT
Epileptogenesis in vivo can be altered by manipulation of molecules such as cytokines and complement that subserve intercellular signaling in both the inflammatory and central nervous systems. Because of the dual roles of these signaling molecules, it has been difficult to precisely define the role of systemic inflammation in epileptogenesis. Organotypic hippocampal brain slices can be maintained in culture independently of the systemic inflammatory system, and the rapid course of epileptogenesis in these cultures supports the idea that inflammation is not necessary for epilepsy. However, this preparation still retains key cellular inflammatory mediators. Here, we found that rodent hippocampal organotypic slice cultures depleted of T lymphocytes and microglia developed epileptic activity at essentially the same rate and to similar degrees of severity as matched control slice cultures. These data support the idea that although the inflammatory system, neurons, and glia share key intercellular signaling molecules, neither systemic nor CNS-specific cellular elements of the immune and inflammatory systems are necessary components of epileptogenesis.

No MeSH data available.


Related in: MedlinePlus

Elimination of microglia using liposomal (Lipo) clodronate from organotypic hippocampal slices of rat. A, Iba-1-positive cells (microglia and macrophages) in CA1 were depleted by using liposomal clodronate (0.2 mg/ml). B, Treatment for 3 d (from DIV9 to 12) decreased the number of microglia comparing liposome-control group and saline control group. Six day treatment from DIV16 to DIV22 eliminated more Iba-1(+) cells than 3 d treatment (96.2% vs 70.4%, n = 4 per group, p = 0.07). C, Microglial depletion persisted 16 d after washout. All values are expressed as mean ± SEM. *p < 0.05. Scale bar, 100 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4596089&req=5

Figure 1: Elimination of microglia using liposomal (Lipo) clodronate from organotypic hippocampal slices of rat. A, Iba-1-positive cells (microglia and macrophages) in CA1 were depleted by using liposomal clodronate (0.2 mg/ml). B, Treatment for 3 d (from DIV9 to 12) decreased the number of microglia comparing liposome-control group and saline control group. Six day treatment from DIV16 to DIV22 eliminated more Iba-1(+) cells than 3 d treatment (96.2% vs 70.4%, n = 4 per group, p = 0.07). C, Microglial depletion persisted 16 d after washout. All values are expressed as mean ± SEM. *p < 0.05. Scale bar, 100 µm.

Mentions: The concentration and duration of clodronate treatment in organotypic slices was optimized to obtain maximal microglial depletion. An equal concentration of empty liposomes was used as the control condition in all experiments utilizing clodronate. When clodronate (final concentration of 0.2 mg/ml in the culture media) was applied for 3 d to rat slices from DIV16 to DIV19, the number of microglia counted in the CA1 subfield decreased by 70.4% (95.5 ± 10.9 vs 28.25 ± 9.8 per field; n = 4; p = 0.004a; Table 2; Fig. 1B). The remaining microglia were small and rounded in shape, and had few ramified processes (Fig. 1A). To increase microglial depletion, the duration of treatment was extended up to 6 d (DIV16−DIV22). This reduced microglia by 96.2% (91.8 ± 15.5 vs 3.5 ± 3.2; n = 4; p = 0.001b; Fig. 1B) compared to controls. The number of Iba-1(−) cells was not different between groups (320.3 ± 8.6 per field in clodronate group vs 325.5 ± 10.3 per field in control group, n = 4, p = 0.77c). The number of CA1 microglia in control slices treated with empty liposomes was not different from slices treated with an equal volume of PBS. Following clodronate depletion, microglia were not regenerated even after 3 weeks in culture (Fig. 1C), indicating that clodronate also effectively eliminated microglial progenitor cells.


What Elements of the Inflammatory System Are Necessary for Epileptogenesis In Vitro?(1,2).

Park KI, Dzhala V, Saponjian Y, Staley KJ - eNeuro (2015)

Elimination of microglia using liposomal (Lipo) clodronate from organotypic hippocampal slices of rat. A, Iba-1-positive cells (microglia and macrophages) in CA1 were depleted by using liposomal clodronate (0.2 mg/ml). B, Treatment for 3 d (from DIV9 to 12) decreased the number of microglia comparing liposome-control group and saline control group. Six day treatment from DIV16 to DIV22 eliminated more Iba-1(+) cells than 3 d treatment (96.2% vs 70.4%, n = 4 per group, p = 0.07). C, Microglial depletion persisted 16 d after washout. All values are expressed as mean ± SEM. *p < 0.05. Scale bar, 100 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4596089&req=5

Figure 1: Elimination of microglia using liposomal (Lipo) clodronate from organotypic hippocampal slices of rat. A, Iba-1-positive cells (microglia and macrophages) in CA1 were depleted by using liposomal clodronate (0.2 mg/ml). B, Treatment for 3 d (from DIV9 to 12) decreased the number of microglia comparing liposome-control group and saline control group. Six day treatment from DIV16 to DIV22 eliminated more Iba-1(+) cells than 3 d treatment (96.2% vs 70.4%, n = 4 per group, p = 0.07). C, Microglial depletion persisted 16 d after washout. All values are expressed as mean ± SEM. *p < 0.05. Scale bar, 100 µm.
Mentions: The concentration and duration of clodronate treatment in organotypic slices was optimized to obtain maximal microglial depletion. An equal concentration of empty liposomes was used as the control condition in all experiments utilizing clodronate. When clodronate (final concentration of 0.2 mg/ml in the culture media) was applied for 3 d to rat slices from DIV16 to DIV19, the number of microglia counted in the CA1 subfield decreased by 70.4% (95.5 ± 10.9 vs 28.25 ± 9.8 per field; n = 4; p = 0.004a; Table 2; Fig. 1B). The remaining microglia were small and rounded in shape, and had few ramified processes (Fig. 1A). To increase microglial depletion, the duration of treatment was extended up to 6 d (DIV16−DIV22). This reduced microglia by 96.2% (91.8 ± 15.5 vs 3.5 ± 3.2; n = 4; p = 0.001b; Fig. 1B) compared to controls. The number of Iba-1(−) cells was not different between groups (320.3 ± 8.6 per field in clodronate group vs 325.5 ± 10.3 per field in control group, n = 4, p = 0.77c). The number of CA1 microglia in control slices treated with empty liposomes was not different from slices treated with an equal volume of PBS. Following clodronate depletion, microglia were not regenerated even after 3 weeks in culture (Fig. 1C), indicating that clodronate also effectively eliminated microglial progenitor cells.

Bottom Line: Organotypic hippocampal brain slices can be maintained in culture independently of the systemic inflammatory system, and the rapid course of epileptogenesis in these cultures supports the idea that inflammation is not necessary for epilepsy.However, this preparation still retains key cellular inflammatory mediators.These data support the idea that although the inflammatory system, neurons, and glia share key intercellular signaling molecules, neither systemic nor CNS-specific cellular elements of the immune and inflammatory systems are necessary components of epileptogenesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurology, Massachusetts General Hospital , Boston, Massachusetts 02129 ; Seoul Paik Hospital, Inje University , Seoul 100-032, South Korea.

ABSTRACT
Epileptogenesis in vivo can be altered by manipulation of molecules such as cytokines and complement that subserve intercellular signaling in both the inflammatory and central nervous systems. Because of the dual roles of these signaling molecules, it has been difficult to precisely define the role of systemic inflammation in epileptogenesis. Organotypic hippocampal brain slices can be maintained in culture independently of the systemic inflammatory system, and the rapid course of epileptogenesis in these cultures supports the idea that inflammation is not necessary for epilepsy. However, this preparation still retains key cellular inflammatory mediators. Here, we found that rodent hippocampal organotypic slice cultures depleted of T lymphocytes and microglia developed epileptic activity at essentially the same rate and to similar degrees of severity as matched control slice cultures. These data support the idea that although the inflammatory system, neurons, and glia share key intercellular signaling molecules, neither systemic nor CNS-specific cellular elements of the immune and inflammatory systems are necessary components of epileptogenesis.

No MeSH data available.


Related in: MedlinePlus