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. 2010 Sep 1;30(35):11745–11761. doi: 10.1523/JNEUROSCI.1769-10.2010

Figure 5.

Figure 5.

The role of NKCC1 in seizure-induced chloride accumulation and facilitation of recurrent seizures. a, Top, Low-Mg2+ ACSF induced recurrent seizures in control and in the presence of NKCC1 blocker bumetanide (10 μm). Extracellular field potential recordings in the CA3 pyramidal cell layer in the intact hippocampus in vitro of neonatal (P5) CLM-1 mouse. Bottom, Example of two-photon confocal imaging of YFP (left) of CA3 neurons yielded pseudocolor map of [Cl−]i before seizures, after two seizures, and after nine seizures in the presence of bumetanide. b, Distribution of [Cl−]i (bin size, 10 mm) in 48 identified neurons before seizures (black), after two seizures in control (low-Mg) solution (red), and after nine seizures in the presence of bumetanide (Bum; green). Gaussian fits yielded mean ± width values of 22.1 ± 19 mm in control, 29.8 ± 18.4 mm after n = 2 seizures, and 44.4 ± 24.1 mm after n = 9 seizures. c, Seizure-induced changes of mean [Cl−]i in subpopulation of neurons with low (0–10 mm), medium (10–20 mm), and high (20–30 mm) range of control resting chloride. Bumetanide prevented seizure-induced neuronal chloride accumulation. d, Effects of recurrent seizures on intracellular chloride accumulation in control low-Mg2+ ACSF and in the presence of bumetanide. Solid lines represent linear regression [Control; low-Mg2+ ACSF (black): r = 0.97 ± 1.66; p = 0.03; bumetanide (green): r = −0.43 ± 2.6; p = 0.57]. In low-Mg2+ ACSF, [Cl−]i progressively increases as a function of recurrent seizures. Bumetanide prevents progressive chloride accumulation induced by recurrent seizures. e, f, Mean interseizure intervals (e) and seizure power (f) as a function of the number of recurrent seizures. e, Solid lines represent linier regression fit of the mean interseizure intervals (ISIs) in control (black; r = −0.82 ± 1.37; p = 0.02) and in the presence of bumetanide (green: r = −0.55 ± 0.7; p = 0.2). f, Corresponding linear regression fit of normalized mean power in control (black; r = 0.95 ± 0.07; p = 0.0003; n = 8) and in the presence of bumetanide (green; r = 0.08 ± 0.08; p = 0.85; n = 8). Bumetanide prevents a progressive increase of seizure power and frequency.