Dendrites of medial olivocochlear neurons in mouse

Abstract

Stains for acetylcholinesterase (AChE) and retrograde labeling with Fluorogold (FG) were used to study olivocochlear neurons and their dendritic patterns in mice. The two methods gave similar results for location and number of somata. The total number of medial olivocochlear (MOC) neurons in the ventral nucleus of the trapezoid body (VNTB) is about 170 per side. An additional dozen large olivocochlear neurons are located in the dorsal periolivary nucleus (DPO). Dendrites of all of these neurons are long and extend in all directions from the cell bodies, a pattern that contrasts with the sharp frequency tuning of their responses. For VNTB neurons, there were greater numbers of dendrites directed medially than laterally and those directed medially were longer (on average, 25-50% longer). Dendrite extensions were most pronounced for neurons located in the rostral portion of the VNTB. When each dendrite from a single neuron was represented as a vector, and all the vectors summed, the result was also skewed toward the medial direction. DPO neurons, however, had more symmetric dendrites that projected into more dorsal parts of the trapezoid body, suggesting that this small group of olivocochlear neurons has very different physiological properties. Dendrites of both types of neurons were somewhat elongated rostrally, about 20% longer than those directed caudally. These results can be interpreted as extensions of dendrites of olivocochlear neurons toward their synaptic inputs: medially to meet crossing fibers from the cochlear nucleus that are part of the MOC reflex pathway, and rostrally to meet descending inputs from higher centers.

Figure 1

Dark AChE staining (black) in presumed OC neurons in the superior olivary complexin a photomicrograph of a transverse section through the left side of the mouse brainstem. Most MOC neurons are in the ventral nucleus of the trapezoid body (VNTB) and extendendrites medially toward the trapezoid body (TB). A few stained neurons are located more dorsally, in the dorsal periolivary nucleus (DPO)and extend dendrites in all directions. Stained MOC axons are visible projecting dorsally, which eventually form the olivocochlear bundle. More lightly-stained presumed LOC neurons are seen within (intrinsic neurons) and on the margins of (shell neurons) the LSO. LOC dendrites and axons are not well stained. Neutral red counterstain. Scale bar: 100 µm.

Figure 2

Medial extensions of dendrites of MOC neurons. Camera lucida drawings of the left-side superior olivary complex, showing (A) all OC somata superimposed from four serial ACh-E stained sections, (B) the large somata and dendrites contained in just one of the sections shown in A, and (C) MOC somata and dendrites from a more rostral section containing only VNTB. In B and C, arrows show the extensive MOC dendrites running medially and dorso-medially toward the trapezoid body (TB). In panel B, DPO neurons and their dendrites face the most dorsal part of the TB. Scale bar: 100 µm.

Figure 3

The similarity in positions of OC neurons retrogradely labeled with Fluorogold (FG, left panels) and stained for AChE (right panels). In these atlases, the individual drawings show the somata within two superimposed 80-µm thick transverse sections; each dot indicates one neuron (except for neurons within the LSO, which are too numerous to be indicated individually). The FG atlas shows the OC neurons retrogradely labeled from a single injected cochlea (on the left side) whereas the AChE atlas shows presumed left-projecting as well as right-projecting OC neurons. The OC bundle of stained axons is indicated on the AChE atlas. VII: facial motor nucleus; DCN, PVCN, AVCN: dorsal, posteroventral, anteroventral cochlear nucleus; DPO: dorsal periolivary nucleus; LSO: lateral superior olive, MNTB: medial nucleus of the trapezoid body; VNTB: ventral nucleus of the trapezoid body. Scale bar: 1 mm. The number of MOC neurons in VNTB (A) is around 150 per side regardless of labeling method, whereas the number of DPO neurons (B) depends on labeling method. Columns show averages and error bars show SEMs. Number of cases counted were AChE: 9 brainstem halves counted in 6 mice; FG: 9 brainstems; HRP: 3 brainstems. FG and HRP counts show numbers on the same (white) and opposite (black) sides of the brainstem relative to the cochlea injected with tracer; AChE counts show number in one half of the brainstem (gray).

Figure 4

Figure 5

Micrograph showing individual MOC neurons with dendrites (arrows) extending medially up to 300 µm as labeled by FG (top) or stained for AChE (bottom). Neurons are located in the rostral VNTB on the left side of the brainstem; medial is toward the right. Scale bar: 100 µm.

Figure 6

Polar plots showing that dendrites are longer in the medial direction for VNTB neurons but not for DPO neurons. Inset: Each dendrite was approximated by a line from soma to tip. Length of this line was used as dendrite length and angle of this line from dorsal was used for dendrite angle. Polar plots show dendrite angles (in degrees) and lengths (in µm) for 152 FG-labeled dendrites (open symbols) from 6 mice brainstems and 136 AChE-stained dendrites (filled symbols) from 2 mice brainstems sectioned in the transverse plane. A: all dendrites from neurons located in VNTB and DPO; B–E: neurons from subgroups separated according to their position. The VNTB caudal group was defined as neurons caudal to the LSO, the middle group as neurons in sections containing the LSO, and the rostral group as neurons rostral to the LSO; these groups had no distinct separation at their boundaries. Numbers of observations of dendrites does not represent the numbers of neurons in each group; for example the group of neurons ventral to the LSO is a dense cluster whose dendrites were intertwined and difficult to separate so fewer points are on this plot (panel B).

Figure 7

Polar plots showing that dendrites are longer in the rostral direction for all neurons. Plots show dendrite angles and lengths for 110 AChE-stained dendrites from 2 mice brainstems sectioned in the sagittal plane. A: all dendrites from neurons located in VNTB and DPO; B–E: neurons from subgroups separated according to their position as described in legend of Fig. 6.

Figure 8

Polar plots showing that the summed path length of all dendrite branches for an individual dendrite is also larger in the medial direction. Data are plotted separately according to section plane for VNTB neurons (A,C) and for DPO neurons (B,D). Shown are data from FG-labeled dendrites (open symbols) and AChE-stained dendrites (filled symbols).

Figure 9

Reconstruction of a single AChE-stained MOC neuron in the VNTB showing preferential extension of the dendrites in the medial direction. All dendrites were contained within the single transverse section (inset, dots show other stained neurons). Part of the axon is also shown. Scale bar: 50 µm. Inset: Position of labeled neuron in the VNTB in the transverse section. Inset scale bar: 0.5 mm.

Figure 10

Reconstruction of a single AChE-stained MOC neuron in the rostral VNTB showing preferential extension of the dendrites in the rostral direction. All dendrites were contained within the single sagittal section. No axon was distinguishable. Scale bar: 50 µm. Inset: Position of labeled neuron in the sagittal section. Inset scale bar: 0.5 mm.

Figure 11

Scatter plot showing dendrite angle to tip (as measured in Fig. 6, inset) is predicted by the dendrite’s initial angle. The initial angle was measured as the dendrite emanates from the cell body (see Methods). Data are from transversely sectioned material including both VNTB and DPO neurons; open symbols plot FG data and closed symbols plot AChE data.

Figure 12

Single neuron vector angles and magnitudes plotted for 17 FG-labeled and 29 AChE-stained neurons that were reconstructed to all their dendrites in the transverse plane. To compute a vector for a single neuron, each of the neuron’s dendrites was treated as a separate individual vector with its own length and angle (Fig. 6, inset), then all vectors of a single neuron were summed. The resultant single neuron vector is plotted on the figure as a point. Points near the origin indicate a pattern of equal-length dendrites radiating symmetrically in all directions; points in the right half of the plot indicate a pattern of more dendrites and/or longer dendrites in the medial direction.