Dendrite Elongation and Dendritic Branching Are Affected Separately by Different Forms of Intrinsic Motoneuron Excitability

Abstract

Duch C, Vonhoff F, Ryglewski S. Dendrite elongation and dendritic branching are affected separately by different forms of intrinsic motoneuron excitability. J Neurophysiol 100: 2525-2536, 2008. First published August 20, 2008 doi:10.1152/jn.90758.2008. Dendrites are the fundamental determinant of neuronal wiring. Consequently dendritic defects are associated with numerous neurological diseases and mental retardation. Neuronal activity can have profound effects on dendritic structure, but the mechanisms controlling distinct aspects of dendritic architecture are not fully understood. We use the Drosophila genetic model system to test the effects of altered intrinsic excitability on postembryonic dendritic architecture development. Targeted dominant negative knock-downs of potassium channel subunits allow for selectively increasing the intrinsic excitability of a selected subset of motoneurons, whereas targeted expression of a genetically modified noninactivating potassium channel decrease intrinsic excitability in vivo. Both manipulations cause significant dendritic overgrowth, but by different mechanisms. Increased excitability causes increased dendritic branch formation, whereas decreased excitability causes increased dendritic branch elongation. Therefore dendritic branching and branch elongation are controlled by separate mechanisms that can be addressed selectively in vivo by different manipulations of neuronal intrinsic excitability.

Preparation of <em>Drosophila</em> Central Neurons for <em>in situ</em> Patch Clamping

“…All other major voltage gated ion channels have been blocked (voltage gated sodium channels with 100 nM tetrototoxin -pipetted directly into the bath; the perfusion was halted for two minutes -calcium channels with 500 μM cadmiumchloride, potassium channels with 2 mM 4-aminopyridin (4-AP) and 30 mM tetraethylammoniumchloride extracellularly and 0.5 mM 4-AP, 20 mM tetraethylammoniumbromide and 144 mM cesiumchloride intracellularly via the intracellular patch solution in the patch pipette (for details see 13,14 ). Action potential traces were evoked by current injection into the MN5 soma (Figure 6) without usage of any ion channel blockers 12 . Figure 1.…”

“…Removal of this sheath is a necessary precondition for patch clamp recordings from any neuron in the adult Drosophila CNS. In recent years scientists have been able to conduct in situ patch clamp recordings from neurons in the adult brain 3,4 and ventral nerve cord of embryonic 5,6 , larval 7,8,9,10 , and adult Drosophila 11,12,13,14 . A stable giga-seal is the main precondition for a good patch and depends on clean contact of the patch pipette with the cell membrane to avoid leak currents.…”

Tiling among stereotyped dendritic branches in an identified Drosophila motoneuron

“…The recombinant C380-GAL4; UAS-mCD8-GFP; Cha-GAL80 line has been described previously (Duch et al, 2008). It expresses predominantly in motoneurons and was obtained from Dr. S. Sanyal (Emory University, Atlanta, GA).…”

“…1A). As is the case with most insect motoneurons, the soma does not generate action potentials but is mostly passive Duch et al, 2008;Meseke et al, 2009). The dendrites of MN5 span large parts of the dorsal mesothoracic motor neuropil and sprout off a primary neurite connecting the cell body with the axon (Fig.…”

“…We conducted branch order analysis by defining the primary neurite as origin, with the branch order 0, as described in previous studies (see Materials and Methods;Evers et al, 2005;Duch et al, 2008;Meseke et al, 2009). This procedure of collapsing the reconstruction onto one virtual origin ensured that all dendrites branching off the primary neurite were considered first-order branches, and the distances and orders of all daughter branches are measured with regard to their branching order relative to this first-order branch.…”