Cochlear innervation in the developing rat: An immunocytochemical study of neurofilament and spectrin proteins

Abstract

We have studied the innervation of the developing cochlea by immunocytochemical staining of the cytoskeletal proteins, neurofilament (NF), and spectrin (brain spectrin and erythrocyte spectrin). NF immunoreactivity was seen in spiral ganglion cell bodies and their processes and in fibers of the intraganglionic spiral bundle (IGSB) on gestational day 16. NF immunoreactivity with monoclonal antibodies to NF160 and NF68 was present beneath both inner hair cells (the IHC) and outer hair cells (OHCs) on gestational day 20. NF200 immunostaining was located only in the IGSB and in fibers reaching the IHC. The first NF200 immunoreactivity beneath the OHCs was seen in the basal turn at birth. NF labelling began to decrease on postnatal day 9 and its intensity became more like that of the adult. Brain spectrin immunostaining was first seen in the IGSB of the basal turn on gestational day 18. It reached the fibers between the spiral ganglion and the IHC on gestational day 20. Brain spectrin immunoreactivity was first seen beneath the OHCs in the basal turn at birth. It reached all the OHCs of the cochlea by postnatal day 4, and began to decrease 9 days after birth. Erythrocyte spectrin immunostaining was first observed during the second postnatal week, when it labelled spiral ganglion cells. The distribution of NF200 and brain spectrin immunoreactivity suggested that efferent innervation of OHCs is present at birth in the rat, and confirms previous studies showing the early efferent innervation of the OHCs of the mouse and the rat at birth, and the time lag between the appearance of the two spectrin isoforms during development.

Expression of Subtypes NF-L, NF-M and NF-H of Neurofilament Triplet Proteins in the Developing Rat Inner Ear

“…The efferent innervation of the OHCs in the embryonic rat forms prenatally, which is earlier than previously believed [10]. Galinovic-Schwartz et al [24] have purified a protein (Mr = 66) of neuronal IFs defined as NF-66 which seems to correspond to a-internexin.…”

“…Moreover, such differences in immunomorphological ex pression (as estimated semiquantitatively) coincided at a time when two populations of differently immunostained spiral and vestibular ganglion cells were already present. Such a difference might show the dual origin of otic gan glion cells, the separation into type I and type II spiral ganglion cells or differences in innervation patterns from Scarpa's ganglion to the two types of vestibular hair cells [9]Immunostaining to each subtypes of NF has been per formed in the developing optic nerve [21], in the develop ing cochlea [10] and in mature spiral ganglion neurons [22]. In developmental neuronal tissue, NF-H is the last NF subtype to be expressed.…”

“…The expression of NFs in the developing inner ear has been studied using different species [4][5][6][7][8][9] including a time sequence analysis of the maturation of the afferent and efferent innervation in the developing rat cochlea [10] and the cytoskeletal differentiation into ganglion cells of type I and type II [10,11], The prerequisite for the onset of inner ear function is a general morphologic maturation, i.e. of hair cells, nerve fibres, synaptogenesis and nerve fibre myelination.…”

Time-dependence and cell-type specificity of synergistic neurotrophin actions on spiral ganglion neurons

“…Enhanced neurofilament content was initially noted with light and electron microscopy (Nishimura et al, 1965;Thomsen, 1966;Kellerhals et al, 1967) and neurofilament stains (Kiang et al, 1982). Subsequent analysis, with immunohistochemical techniques, showed a predominance of neurofilament 68 and 160, phosphorylated neurofilament 200, and peripherin in these same neurons (Dau and Wenthold, 1985;Ryugo, 1986, 1991;Romand et al, 1990;Hafidi et al, 1990Hafidi et al, , 1993Anniko et al, 1995). These markers are useful in vitro because other ways of classifying the cells were not possible.…”

The Structural and Functional Differentiation of Hair Cells in a Lizard's Basilar Papilla Suggests an Operational Principle of Amniote Cochleas

“…After labeling auditory nerve fibers with an antibody against neurofilament-associated protein 200, which is expressed in both afferent and efferent fibers (Hafidi et al, 1990), we used laserscanning confocal microscopy to reconstruct the peripheral trajectories of the axons innervating the hair cells in five cochleas. Because there is no efferent innervation in the high-frequency region of lizard basilar papillas (Manley, 1990), the auditory nerve fibers in this region represent only afferent terminals, the distal axonal segments connecting hair cells to eighth-nerve neurons.…”