It has been proposed that HERG plays a role in spike-frequency adaptation, a property shown by a large number of central and peripheral neurones, in which a gradual increase in K⁺ conductance counteracts the Na⁺ currents and thus leads to either a longer interspike interval or a complete block of the burst (Chiesa et al. 1997). An immuno-EM study of the rat hippocampus using two distinct antibody preparations failed to find erg protein in neurones, although it was detected in astrocytes (Emmi et al. 2000). We performed immuno-histochemistry on horizontal sections of rat brain (humanely killed) and human hippocampus (post-mortem, with consent) using a commercially available anti-HERG antibody (Alomone, APC-016). This antibody was able to detect HERG protein in HERG transiently transfected CHO cells using immunofluorescence; HERG was abundantly detected by this antibody in the cytoplasm of CHO cells, although not localised in the membrane. CHO cells have electrophysiologically detectable HERG currents after transfection, but not without transfection. This antibody stained a small population of neurones in the brain asymmetrically in the cytoplasm. The stained neurones were found in the cortex and regions of the hippocampus (stratum radiatum and stratum lacunosum-moleculare (LM)), but not in the hindbrain. Morphology of the stained cells varied by region, including some archetypal bipolar morphologies in layer 1 of the cortex. To test whether HERG staining cells represent inhibitory interneurones, we alternatively stained numbered serial sections with anti-HERG antibody or anti-GABA antibody (Sigma, A-2052), and found that some HERG staining cells aligned with GABA staining cells in adjacent sections. We used the whole-cell patch-clamp technique to determine whether erg channels could be detected in neurones in layer 5 of the rat entorhinal cortex (EC) and in visually identified inhibitory interneurones in the hippocampus. Voltage- and current-clamp recordings in hippocampal neurones in the radiatum and LM using both physiological and raised K⁺ (10 mM) extracellular solutions failed to demonstrate an E-4031-sensitive HERG tail current or a change in spike frequency adaptation in response to positive current injection. However, analysis revealed an increase in the frequency of spontaneous excitatory postsynaptic currents (EPSCs) in layer 5 of the EC in response to bath application of the HERG-specific blocker E-4031. These data highlight the differences in the ability to detect E-4031-sensitive currents in the two electrophysiological approaches. The immunohistochemistry represents a novel finding of erg protein in neurones in the brain, and suggests that further immunohistochemistry using our methodology with another anti-erg antibody needs to be performed.

Chiesa, N., Rosati, B., Arcangeli, A., Olivotto, M. & Wanke, E. (1997). J. Physiol. 501, 313-318. abstract

Emmi, A., Wenzel, H.J., Schwartzkroin, P.A., Taglialatela, M., Castaldo, P., Bianchi, L., Nerbonne, J., Robertson, G.A. & Janigro, D. (2000). J. Neurosci. 20, 3915-3925.