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Exp Brain Res (2003) 153: 514–521 DOI 10.1007/s00221-003-1617-z

RESEARCH ARTICLE

D. O. Kim . X. M. Yang . Y. Ye A subpopulation of neurons retrogradely labeled with cholera toxin-B injected into the inner ear

Received: 20 December 2002 / Accepted: 2 July 2003 / Published online: 5 September 2003 # Springer-Verlag 2003

Abstract Previous studies have shown that: (1) raphe vestibular multisensory event, the raphe projections to the neurons respond to acoustic and vestibular stimuli, some inner ear and other auditory and vestibular structures may , with a latency of 10–15 ms; (2) alterations of the raphe enhance the mammal s ability to localize and recognize the nuclei alter the acoustic startle reflex; (3) the dorsal raphe sound and respond properly. The raphe-labyrinthine , nucleus (DRN) is the major source of serotonergic projection may also modulate the inner ear s sensitivity neurons; and (4) approximately 57% of the DRN neurons as a function of the sleep–wake arousal state of an are nonserotonergic. In the present study, cholera toxin organism on a slower time course. subunit-B (CTB) was injected into cat cochleas, and the brain tissue was examined after a survival period of 5– Keywords Auditory . Vestibular . Efferent . Multisensory . 7 days. Aside from neurons which were known to project Monoaminergic . Cat to the inner ear, i.e., olivocochlear and vestibular efferent neurons, a surprising new finding was made that somata of a subpopulation of DRN neurons were intensely labeled Introduction with CTB. These CTB-labeled neurons were densely distributed in a dorsomedian part of the DRN with some in The form a rather narrow, more or less a surrounding area outside the DRN. The present results continuous collection of neurons around the midline of the suggest that a novel raphe-labyrinthine projection may that extend from the caudal end of the medulla exist. A future study of anterograde labeling with to the rostral (Taber et al. 1960). The dorsal injections of a tracer in the DRN will be needed to raphe nucleus (DRN), one of eight raphe nuclei which establish the existence of a raphe-labyrinthine projection have been identified on the basis of their location and more thoroughly. A raphe-labyrinthine descending input, cytoarchitecture, is located mostly in the midbrain. The together with an ascending input from the inner ear to the raphe nuclei are the principal sites of the brain for DRN through intervening neurons, such as the juxta- serotonergic neurons (Dahlstrom and Fuxe 1964; Charara acousticofloccular raphe neurons (JAFRNs) described by and Parent 1998) and contain approximately 78% of all Ye and Kim, may mediate a brain stem reflex whereby a serotonergic neurons of the cat brain stem (Wiklund et al. salient multisensory (including auditory and vestibular) 1981). Among the raphe nuclei, the DRN is the major stimulus may alter the sensitivity of the inner ear. As a source of serotonergic neurons and contains approximately mammal responds to a biologically important auditory- 40% of all serotonergic neurons (Wiklund et al. 1981). Even though the DRN is the major source of serotonergic *) . . D. O. Kim ( X. M. Yang Y. Ye neurons of the brain, approximately 57% of DRN neurons Department Neuroscience, University of Connecticut Health Center, are nonserotonergic (Wiklund et al. 1981). The projections Farmington, CT 06030, USA from and to cell groups of the raphe nuclei have been e-mail: [email protected] extensively studied with a variety of techniques and animal species (Brodal 1981; Nieuwenhuys 1985; Vertes X. M. Yang Department of Otolaryngology, Xiang Ya Second Hospital, 1991; Jacobs and Azmitia 1992; Allen and Cechetto 1994; Central South University, Simpson et al. 1998; Morin and Myer-Bernstein 1999). Changsha, Hunan, China Physiologically, many DRN neurons respond to sounds (Le Moal and Olds 1979; Heym et al. 1982; Rasmussen et Present address: – Y. Ye al. 1986), some with a latency of 10 15 ms (Le Moal and Department Surgery, Creighton University School of Medicine, Olds 1979), and neurons in caudal medullary raphe nuclei Omaha, Neb., USA respond to vestibular stimuli with a latency of 10–15 ms

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515 (Yates et al. 1992, 1993). A possible pathway for auditory unexpectedly in a study where CTB was injected into and vestibular signals to reach the DRN is through the the cochlea with the intention of labeling olivocochlear juxta-acousticofloccular raphe neurons (JAFRNs; Ye and neurons retrogradely. To our knowledge, a direct projec- Kim 2001), which are embedded in fiber fascicles just tion from the DRN to the inner ear has not been previously

outside the near the flocculus, restiform reported. Results of this study were presented at the

, body, and . The JAFRNs were found with , Central Auditory Processing—Integration of Auditory ,, retrograde labeling of neurons following injection of and Nonauditory Information meeting, Ascona, Switzer- cholera toxin subunit-B (CTB) into the DRN. The land, May 2002. JAFRNs are well poised to receive multisensory inputs from the cochlear nucleus, flocculus, and vestibular nuclei and provide the combined multisensory information to the Materials and methods DRN. The DRN was also observed to receive inputs from vestibular nuclei (Kawasaki and Sato 1981; Kalen et al. General procedures 1985). Many parts of the mammalian auditory pathways The main results of this study were obtained from four adult cats. Results from three additional adult cats served as controls (described spanning from the to the auditory cortex are below). Each cat weighed 2.0–3.5 kg and was purchased from a innervated by serotonergic neurons (Willard et al. 1984; commercial breeder of research cats (Liberty Research, Waverly, N. Campbell et al. 1987; Klepper and Herbert 1991; Y.). The cats had clean ear canals, no middle-ear infection, and no Thompson et al. 1994, 1995, 1998; Thompson and known hereditary abnormalities. The procedures used in this study followed the Public Health Service Policy on Humane Care and Use Thompson 1995) or by neurons of the raphe nuclei of Laboratory Animals, and were approved by the Animal Care (Adams and Warr 1976; Klepper and Herbert 1991; Committee, University of Connecticut Health Center. Each cat was Thompson et al. 1995). Among the various raphe nuclei, anesthetized with an intraperitoneal injection of pentobarbital the DRN contains the largest number of cells that project sodium, 35 mg/kg. Deep anesthesia was maintained by administer- ing pentobarbital sodium through an intravenous catheter. Surgical, to brain stem auditory nuclei (Klepper and Herbert 1991; procedures were carried out under sterile conditions. Each animal s Thompson et al. 1995). The raphe nuclei, including the body temperature was maintained near 38°C with a regulated DRN, also contain many nonserotonergic neurons (Wik- heating pad. lund et al. 1981; Charara and Parent 1998). The relation- In the main experiment, the bulla was opened to visualize the ship between serotonergic and nonserotonergic neurons of round window membrane. A micropipette with an inner diameter of 40–70 μm was advanced with a micromanipulator to impale the the DRN and their innervation of auditory brain stem round window membrane. The micropipette was filled with a 1% structures has not been well described in the literature. solution of CTB (List Biological Laboratories, Campbell, Calif.). A Neurons of vestibular nuclei also have serotonergic volume of 3–6 μl CTB was injected into the scala tympani of one innervation (Steinbusch 1981), and are influenced by cochlea in each cat through the micropipette using a pressure injector (Picospritzer; General Valve, East Hanover, N.J.) over a activation of the DRN, some mediated by serotonergic period of several minutes. When the micropipette was withdrawn inputs (Kishimoto et al. 1991; Licata et al. 1995) and under examination with an operating microscope, there was no others mediated by nonserotonergic inputs (Licata et al. visible leakage of fluid from the round window. To prevent any 1995). leakage of the tracer through the penetration point, a piece of connective tissue from the neck muscle of the cat was placed on the Considering that the DRN is generally involved in membrane so as to seal the round window. modulating the nervous system as a function of a , In one cat, a control experiment was performed as follows. On the mammal s behavioral state (Trulson and Jacobs 1979; left side, the cerebellum was exposed and retracted, exposing the Jacobs and Azmitia 1992), the above observations suggest cochlear nucleus. A volume of 14 μl CTB was injected into a pool of that the DRN may be able to combine information from cerebrospinal fluid (CSF) surrounding the cochlear nucleus. The craniotomy was closed by suturing overlying muscles. Results from multiple sensory systems with information about the two other cats served as the second type of control. In these two cats, behavioral state of a mammal. In this context, the the experimental procedure was the same as that of the main projections from the DRN to auditory and vestibular experiment. Details about the latter two cats are described in the structures may underlie modulation of the auditory and Results section. vestibular systems, reflecting both the behavioral state of a mammal and the environmental state conveyed by mul- Histological processing tiple sensory systems. The cochlea and vestibular end organs of the inner ear After a survival period of 5–7 days, each cat was anesthetized and are innervated by olivocochlear and vestibular efferent perfused transcardially with saline, followed by a fixative solution neurons, respectively (Warr 1992; Dechesne et al. 1984). containing 4% paraformaldehyde in 0.1 M sodium phosphate buffer, These neurons, which are located in the , form brain pH 7.4, and 0.004% calcium chloride. The survival period of the cat with CTB injection into the CSF was 5 days. The brain stem was stem reflex arcs involving the inner ear. Besides these blocked using a stereotaxic frame (David Kopf Instruments, pontine efferent labyrinthine neurons, no other brain stem Tujunga, Calif.). neurons are known to innervate the inner ear. The present The brain tissue was kept in 30% sucrose for 12–24 h. Frozen paper describes a surprising finding of novel, putative sections were cut at 30–50 μm in the transverse plane. The brain raphe-labyrinthine projection from the DRN to the inner sections were rinsed in 10 mM phosphate-buffered saline (PBS) containing 0.9% sodium chloride, 10 min ×3, between each of the ear. The DRN is located far rostral to the olivocochlear and following steps. The sections were incubated with 5% normal vestibular efferent neurons. This finding was made 中国科技论文在线 http://www.paper.edu.cn

516 donkey serum (Jackson Immuno Research Laboratories, West Grove, Pa.), 1 h, and then in a primary antibody (goat anti-CTB; List Biological Laboratories, Campbell, Calif.), diluted to 1:5,000 in 10 mM PBS, plus 0.2% triton X-100, 1 day, 4°C, under constant agitation. The sections were then incubated in a secondary antibody (donkey anti-goat IgG; Jackson Immuno Research Laboratories), diluted to 1:500 in 10 mM PBS, plus 0.2% triton X-100, 2 h, room temperature. The secondary antibody was conjugated with fluores- cent label, Texas Red. The sections were mounted on glass slides with Vectashield (Vector Laboratories, Burlingame, Calif.) and cover-slipped.

Microscopic examination

A light microscope (Axioplan; Carl Zeiss, Thornwood, N.Y.) was used to examine the neurons labeled with CTB. The microscope was equipped with a mercury-arc light source and a set of filters for fluorescent microscopy. Cells labeled with CTB/Texas-Red were visualized with a red filter. Digital images of the sections were captured with a camera (Spot Insight color camera; Micro Video Instruments, Avon, Mass.) connected to a PC running Windows software. The images were processed with Photoshop (Adobe Systems) and Corel Draw (Corel, Ottawa, Canada) programs.

Results

The location of the DRN is shown in Fig. 1. Figure 1A and B represent midsagittal and transverse sections of a cat brain stem, respectively. The DRN is located in caudal midbrain in a dorsal region around the midline. A small square box around a dorsomedian part of the DRN in Fig. 1B corresponds to the areas represented in Figs. 2 and 3. Images of DRN neurons retrogradely labeled with CTB from one cat are shown in Fig. 2. The figure represents a transverse section of the DRN with the dorsal side up. The midline of the brain stem is an approximate vertical line Fig. 1A, B Drawings of the cat brain stem describing the location running near the center in Fig. 2A, B. Figure 2B is a of the dorsal raphe nucleus (DRN). A Sagittal plane near the magnified version of the box in A. Figure 2C and D are midline; B transverse plane. A small square box around a magnified versions of the two boxes in B. Note that dorsomedian part of the DRN in B corresponds to the fields Fig. 2C and D were taken at different focal planes. The shown in Figs. 2A and 3A. (A anterior, AQ aqueduct, BP brachium dorsal surface of the brain stem (i.e., the floor of the 4th pontis, cerebel. cerebellum, D dorsal, DNLL dorsal nucleus of , DRN dorsal raphe nucleus, IC inferior colliculus, IC-Co ventricle) is indicated by a dashed line at the top of commissure of inferior colliculus, IP interpeduncular nucleus, L Fig. 2A. In Fig. 2A, B, the DRN is densely packed with lateral, LL lateral lemniscus, MLF medial longitudinal fasciculus, labeled neurons, which are concentrated in a dorsomedian MRN median raphe nucleus, OMN oculomotor nucleus, P pyramidal part of the DRN. The ventral part of the DRN and the tract, PAG periaqueductal gray, SC superior colliculus, TB trapezoid medial longitudinal fasciculi are below the field shown in body, 4th Vent. fourth ventricle) Fig. 2A. In the ventral part of the DRN, there were only a few scattered labeled neurons. These DRN neurons were had other differences besides their soma size. The small , intensely labeled with CTB, and the radiating dendrites of neurons somata were rounder, their dendritic trees were some neurons are clearly visible in Fig. 2. less prominent, and their CTB labeling was less intense , In Fig. 2C, D, further details of the cells somata and (but clearly above the background) than those of the large dendrites are visible. For example, branching of dendrites neurons. of a large neuron is well visible near the center in Fig. 2C; Figure 3 shows CTB-labeled neurons in the DRN of the equivalent diameter of this neuron (i.e., the diameter of another cat. The format of Fig. 3 is the same as that of a circle matching the soma area) was 25 μm. The Fig. 2. The results in Fig. 3 are consistent with those of equivalent diameters of other large labeled neurons were Fig. 2 in that cells in the DRN were intensely labeled with 23–27 μm. In Fig. 2D, several small neurons were in CTB and that the labeled cells were concentrated in a focus. The diameters of their somata were 15–19 μm. dorsomedian part of the DRN. It is also seen in Fig. 3 that When compared with the large neurons, the small neurons there were large (21–24 μm diameter) and small (15– 中国科技论文在线 http://www.paper.edu.cn

517 Fig. 2A-D Neurons in a dorso- median part of the DRN that were retrogradely labeled with cholera toxin subunit-B (CTB) injected into the left cochlea in a cat. CTB label was visualized using secondary antibody con- jugated with Texas Red, and these images were taken with fluorescent microscopy. In A, the floor of the fourth ventricle is marked by a dashed line near the top of the panel; the midline runs vertically near the center of the panel, and the field shown is dorsal to the MLF. B A magni- fied image of the area indicated by a box in A. Analogously, C and D represent the lower left and upper right boxes indicated in B, respectively. C and D were taken at different focal planes

18 μm diameter) labeled neurons in the DRN. As in the intensely labeled with CTB. Considering that the amount , previous figure, the small neurons somata were rounder, of CTB injected into the CSF was about 3 times as large as their dendritic trees were less prominent, and their CTB that into the cochlea, i.e., 14 μl versus 3–6 μl, there should labeling was less intense than those of the large neurons. have been more intense labeling of DRN neurons in this Among the CTB-labeled neurons, some of the somata had control cat than in the cats of the main experimental group long and short axes. The long axes were lateromedial (e.g., if the main results had been an artifact of CTB leakage cells in the upper part of Fig. 3C), dorsoventral (e.g., the from the cochlea into the CSF through the cochlear top left cell in Fig. 2A), or oblique (e.g., the top right cell aqueduct. Because the control results were contrary to this in Fig. 2A). prediction, it is suggested that the main results of this By sampling a subset of the total sections, it was study may not represent an artifact of CTB leakage from estimated that there were approximately 1,000 neurons the cochlea into the CSF. On the other hand, the presence labeled with CTB in the vicinity of the DRN in each cat. of faint labeling of DRN neurons after injection of CTB The CTB-labeled neurons were mostly confined within the into the CSF suggests that, at least, some of the main DRN, with a small number of neurons scattered at the results of this study may represent leakage of CTB from margins of the medial longitudinal fasciculus, in the the cochlea into the CSF through the cochlear aqueduct. periaqueductal gray, and in the median raphe nucleus. In two cats (serving as the second type of control), there Along the rostrocaudal axis, the density of labeled neurons was evidence that CTB leaked into the middle ear in that was maximum in the middle of the DRN and it declined CTB-labeled neurons included not only olivocochlear toward both rostral and caudal directions. High intensity of neurons and vestibular efferent neurons, but also motor CTB retrograde labeling and the spatial pattern of labeled neurons of the middle-ear muscles described in the neurons in the DRN region were consistent in all four cats literature, i.e., neurons associated with the motor trigem- of the main experimental group. inal nucleus innervating the tensor tympani muscle and In the cat where CTB was injected into the CSF neurons associated with the facial nucleus innervating the surrounding the cochlear nucleus, there was only faint stapedius muscle. The number of CTB-labeled olivoco- labeling of DRN neurons, without any of them being chlear neurons in the present study was comparable with 中国科技论文在线 http://www.paper.edu.cn

518 Fig. 3A-D Neurons in a dorso- median part of the DRN that were retrogradely labeled with CTB injected into the left co- chlea in another cat. Similar format to Fig. 2

what was described in the literature (Arnesen and Osen As stated in the Introduction, approximately 57% of 1984; Warren and Liberman 1989; Warr et al. 2002). In the DRN neurons are nonserotonergic (Wiklund et al. 1981). four cats of the main experimental group, there was no These neurons contain such as norepi- such labeling of motor neurons of the middle-ear muscles, nephrine, dopamine, gamma-aminobutyric acid (GABA), indicating an absence of leakage of CTB into the middle and various neuropeptides (Wiklund et al. 1981; Charara ear. Neurons of the DRN were intensely labeled with CTB and Parent 1998). Which (s) the putative regardless of whether CTB leaked into the middle ear. raphe-labyrinthine neurons contain is presently unknown. Therefore, it is concluded that the main results of this study do not represent an artifact of CTB leakage from the cochlea into the middle ear. Transneuronal labeling?

The intensity of transneuronal labeling is expected to be Discussion fainter than that of direct retrograde labeling, because a smaller amount of a tracer is expected in secondary The main finding of this study is the identification of neurons than in neurons that directly take up the tracer. , putative raphe-labyrinthine neurons in the DRN by using The intensity of CTB labeling of DRN neurons somata in retrograde labeling with injection of CTB into the cochlea. the present study (Figs. 2 and 3) is high and quite similar The only efferent labyrinthine neurons in the brain stem, to those of olivocochlear and vestibular efferent neurons that were previously described in the literature, are obtained using the same procedure in our laboratory (Ye at olivocochlear and vestibular efferent neurons, both located al. 2000; Warr et al. 2002). If CTB were transneuronally in the hindbrain. Firm establishment of the existence of a transported, one should see CTB-labeled neurons in raphe-labyrinthine projection will require demonstration of auditory and vestibular structures such as the cochlear anterograde labeling of nerve fibers in the inner ear with a nucleus and vestibular nuclei, which receive projections tracer injected into the DRN. from the cochlear and vestibular ganglion cells. We observed that the primary cochlear and vestibular nerve 中国科技论文在线 http://www.paper.edu.cn

519 , fibers endings were labeled with CTB, but the somata of 1992) including: (1) modulation of sensory systems the cochlear nucleus and vestibular nuclei neurons were including the auditory system (Ebert and Ostwald 1992; free of CTB label. These observations support our view Marriage and Barnes 1995), vestibular system (Kishimoto that the CTB labeling of DRN neurons represents a direct et al. 1991; Licata et al. 1995), visual system (Villar et al. retrograde, rather than a transneuronal, transport. 1988; Waterhouse et al. 1993), and nociceptive system (Basbaum and Fields 1984; Fields et al. 1991; Leung and Mason 1999); (2) modulation of motor function including Target of the raphe-labyrinthine projection: cochlea motor neurons of the spinal cord (White and Fung 1989), and/or vestibular end organ. trigeminal-facial nuclei (Li et al. 1993; including middle- ear-muscle motor neurons, Thompson et al. 1998), and Even though we did not inject CTB directly into the oculomotor nuclei (Buttner-Ennever et al. 1988); (3) vestibular end organ, CTB is believed to have diffused into modulation of brain stem reflexes such as the acoustic the vestibular end organ because, besides olivocochlear startle reflex (Davis and Sheard 1974; Davis et al. 1980; neurons, vestibular efferent neurons (similar to those Rigdon and Weatherspoon 1992; Rasmussen et al. 1997); described by Warr 1975; Dechesne et al. 1984) were also (4) regulation of a sleep–wake arousal state (Puizillout et labeled with CTB. Therefore, we cannot determine al. 1981); and (5) modulation of autonomic function (van whether the cochlea, the vestibular end organs or both de Kar and Lorens 1979; Leung and Mason 1996). receive(s) the raphe-labyrinthine projection. By using an The putative raphe-labyrinthine projection described in antibody against , Gil-Loyzaga et al. (1997) have the present study extends the possible ways whereby the reported that serotonin-immunoreactive fibers are present raphe neurons may modulate the auditory and vestibular in the cat cochlea. The origin of these fibers were not systems. The present finding of the putative raphe- determined and the authors conjectured that the fibers may labyrinthine projection together with ascending inputs to belong to lateral olivocochlear fibers. Whether these the DRN from the auditory (Le Moal and Olds 1979; Ye serotonin-immunoreactive fibers originate from the DRN and Kim 2001) and vestibular systems (Kawasaki and Sato is presently unknown. 1981; Kalen et al. 1985; Yates et al. 1992, 1993; Ye and Kim 2001 ) suggests that reflexes involving the inner ear and the DRN may be produced. The presence of raphe Why may the raphe-labyrinthine projection have been innervation of multiple auditory structures, i.e., cochlear missed previously? nucleus, motor neurons of the middle-ear muscle, olivocochlear neurons, inferior colliculus, and the auditory Potential answers to this question are as follows: First, a cortex (references cited above) implies that raphe nuclei high sensitivity of CTB may have facilitated the present can modulate the auditory system at multiple levels. positive finding. CTB is considerably more sensitive (Wan Analogously, the raphe nuclei may modulate the vestibular et al. 1982; Trojanowski et al. 1982; Vetter and Mugnaini system at the level of vestibular nuclei (Kishimoto et al. 1992; Ye at al. 2000) than horseradish peroxidase (HRP), 1991; Licata et al. 1995) and end organs. widely used previously. CTB may have a high affinity to a As a mammal responds to a biologically important certain population of neurons such that a neuroanatomical multisensory event, e.g., head turning toward a sound projection may be well detectable with CTB but not with signaling a danger, the raphe projections to the inner ear other tracers. Such an example was reported in a recent and other auditory and vestibular structures may enhance , study by Marsh et al. (2002), where a projection from the the mammal s ability to localize and recognize the sound amygdala to the inferior colliculus was much more clearly and respond properly, e.g., escaping from danger signaled detected with CTB than with Fluoro-Gold, Fluororuby, or by the sound. The raphe-labyrinthine projection may also , wheat germ-agglutinated HRP. modulate the inner ear s sensitivity as a function of the Second, the DRN is considerably more rostral to the sleep–wake arousal state of an organism on a slower time olivocochlear and vestibular efferent neurons. Therefore, course. studies with injections of retrograde tracers into the inner ear may have often excluded the portion of the brain stem Acknowledgements This study was supported by NIDCD-NIH

that includes the DRN in histological processing. Often, grant DC00360. We thank D. Machado for making comments on the , , ,, the brain is trimmed to a relevant portion before manuscript. subjecting it to a lengthy and costly sequence of histological procedures. This was the case in the present study until we serendipitously included a more rostral part References of the brain stem, which led us to the present finding. , Adams JC, Warr WB (1976) Origins of axons in the cat s acoustic striae determined by injection of horseradish peroxidase into severed tracts. J Comp Neurol 170:107–122 Possible functions of the raphe-labyrinthine projection Allen GF, Cechetto DF (1994) Serotoninergic and nonserotoninergic neurons in the medullary raphe system have axon collateral The raphe neurons and serotonin have been implicated in a projections to autonomic and somatic cell groups in the medulla and spinal cord. J Comp Neurol 350:357-366 variety of functions (reviewed by Jacobs and Azmitia 中国科技论文在线 http://www.paper.edu.cn

520 Arnesen AR, Osen KK (1984) Fibre population of the vestibuloco- Marriage J, Barnes NM (1995) Is central hyperacusis a symptom of chlear anastomosis in the cat. Acta Otolaryngol (Stockh) 5-hydroxytryptamine (5-HT) dysfunction? J Laryngol Otol 98:255–269 109:915–921 Basbaum AI, Fields HL (1984) Endogenous pain control systems: Marsh RA, Fuzzessery ZM, Grose CD, Wenstrup JJ (2002) brainstem spinal pathways and endorphin circuitry. Ann Rev Projection to the inferior colliculus from the basal nucleus of Neurosci 7:309–338 the amygdala. J Neurosci 22:10449–10460 Brodal A (1981) Neurological anatomy in relation to clinical Morin LP, Meyer-Bernstein EL (1999) The ascending serotonergic medicine. Oxford University Press, New York system in the hamster: comparison with projections of the Buttner-Ennever JA, Cohen B, Pause M, Fries W (1988) Raphe dorsal and median raphe nuclei. Neuroscience 91:81-105 nucleus of the pons containing omnipause neurons of the Nieuwenhuys R (1985) Chemoarchitecture of the brain. Springer, oculomotor system in the monkey, and its homologue in man. J Berlin Comp Neurol 267:307-321 Puizillout JJ, Gaudin-Chazal G, Sayadi A, Vigier D (1981) Campbell MJ, Lewis DA, Foote SL, Morrison JH (1987) Distribu- Serotoninergic mechanisms and sleep. J Physiol (Paris) tion of choline acetyltransferase-, serotonin-, dopamine beta- 77:415-424 hydroxylase-, tyrosine hydroxylase-immunoreactive axons in Rasmussen K, Strecker R, Jacobs BL (1986) Single unit response of monkey primary auditory cortex. J Comp Neurol 261:209-220 noradrenergic, serotonergic and dopaminergic neurons in freely Charara A, Parent A (1998) Chemoarchitecture of the primate dorsal moving cats to simple sensory stimuli. Brain Res 369:336–340 raphe nucleus. J Chem Neuroanat 72:111–127 Rasmussen K, Kallman MJ, Helton DR (1997) Serotonin-1A Dahlstrom A, Fuxe K (1964) Evidence for the existence of antagonists attenuate the effect of nicotine withdrawal on the monoamine-containing neurons in the central nervous system. auditory startle response. Synapse 27:145–152 I. Demonstration of monoamines in the cell bodies of the brain Rigdon GC, Weatherspoon JK (1992) 5-Hydroxytryptamine-1a stem neurons. Acta Physiol Scand (Suppl 232) 62:1–55 receptor agonists block repulse inhibition of acoustic startle Davis M, Sheard MH (1974) Habituation and sensitization of the rat reflex. J Pharmacol Exp Ther 263:486–493 startle response: effect of raphe lesions. Physiol Behav 12:425– Simpson KL, Fisher TM, Waterhouse BD, Lin RC (1998) Projection 431 patterns from the raphe nuclear complex to the ependymal wall Davis M, Astrachan DI, Kass E (1980) Excitatory and inhibitory of the ventricular system in the rat. J Comp Neurol 399:61-72 effects of serotonin on sensorimotor reactivity measured with Steinbusch HWM (1981) Distribution of serotonin-immunoreactiv- acoustic startle. Science 209:521–523 ity in the central nervous system of the rat—cell bodies and Dechesne C, Raymond J, Sans A (1984) The efferent vestibular terminals. Neuroscience 6:557–618 system in the cat: a horseradish peroxidase and fluorescent Taber E, Brodal A, Walberg F (1960) The raphe nuclei of the brain retrograde tracers study. Neuroscience 11:893–901 stem in the cat. I. Normal topography and cytoarchitecture and Ebert U, Ostwald J (1992) Serotonin modulates auditory informa- general discussion. J Comp Neurol 114:161–187 tion processing in the cochlear nucleus of the rat. Neurosci Lett Thompson AM, Thompson GC (1995) Light microscopic evidence 145:51–54 of serotoninergic projections to olivocochlear neurons in the Fields HL, Heinricher MM, Mason P (1991) Neurotransmitters in bushy baby (Otolemur garnettii). Brain Res 695:263–266 nociceptive modulatory circuits. Annu Rev Neurosci 14:219– Thompson GC, Thompson AM, Garret KA, Britton BH (1994) 245 Serotonin and serotonin receptors in the central auditory Gil-Loyzaga P, Bartolome MV, Vicente-Torres MA (1997) Seroto- system. Otolaryngol Head Neck Surg 110:93–102 nergic innervation of the organ of Corti of the cat cochlea. Thompson AM, Moore KR, Thompson GC (1995) Distribution and Neuroreport 8:3519–3522 origin of serotoninergic afferents to guinea pig cochlear Heym J, Trulson ME, Jacobs BL (1982) Raphe unit activity in freely nucleus. J Comp Neurol 351:104–116 moving cats: effects of phasic auditory and visual stimuli. Brain Thompson AM, Thompson GC, Britton BH (1998) Serotoninergic Res 232:29–39 innervation of stapedial and tensor tympani motoneurons. Brain Jacobs BL, Azmitia EC (1992) Structure and function of the brain Res 787:175–178 serotonin system. Physiol Rev 72:165–229 Trojanowski JQ, Gonatas JO, Gonatas NK (1982) Horseradish Kalen P, Karlson M, Wiklund L (1985) Possible excitatory amino peroxidase (HRP) conjugates of cholera toxin and lectins are acid afferents to nucleus raphe dorsalis of the rat investigated more sensitive retrogradely transported markers than the free 3 with retrograde wheat germ agglutinin and D-[ H]aspartate HRP. Brain Res 231:33–50 tracing. Brain Res 360:285–297 Trulson ME, Jacobs BL (1979) Raphe unit activity in free moving Kawasaki T, Sato Y (1981) Afferent projections to the caudal part of cats: correlation with level of behavioral arousal. Brian Res the dorsal nucleus of the raphe in cats. Brain Res 211:439–444 163:135–150 Kishimoto T, Sasa M, Takaori S (1991) Inhibition of lateral van de Kar LD, Lorens SA (1979) Differential serotonergic vestibular nucleus neurons by 5-hydroxytryptamine derived innervation of individual hypothalamic nuclei and other from the dorsal raphe nucleus. Brain Res 553:229–237 forebrain regions by the dorsal and median midbrain raphe Klepper A, Herbert H (1991) Distribution and origin of noradren- nuclei. Brain Res162:45-54 ergic and serotonergic axons in the cochlear nucleus and Vertes PR (1991) A PHA-L analysis of ascending projections of the inferior colliculus of the rat. Brain Res 557:190–201 dorsal raphe nucleus in the rat. J Comp Neurol 313:643–668 Le Moal M, Olds M (1979) Unit responses to auditory input in the Vetter DE, Mugnaini E (1992) Distribution and dendritic features of dorsal and median raphe nuclei of the rat. Physiol Behav three groups of rat olivocochlear neurons. A study with two 22:11–15 retrograde cholera toxin tracers. Anat Embryol (Berl) 185:1-16 Leung CG, Mason P (1996) Spectral analysis of arterial blood Villar MJ, Vitale ML, Hokfelt T, Verhofstad AA (1988) Dorsal pressure and raphe magnus neuronal activity in anesthetized raphe serotoninergic branching neurons projecting both to the rats. Am J Physiol 271:483-489 lateral geniculate body and superior colliculus: a combine Leung CG, Mason P (1999) Physiological properties of raphe retrograde tracing-immunohistochemical study in the rat. J magnus neurons during sleep and waking. J Neurophysiol Comp Neurol 277:126–140 81:584-595 Wan XC, Trojanowski JQ, Gonatas JO (1982) Cholera toxin and Licata F, Li Volsi G, Maugeri G, Sananagelo F (1995) Neuronal wheat germ agglutinin conjugates as neuroanatomical probes: responses in vestibular nuclei to dorsal raphe electrical their uptake and clearance, transganglionic and retrograde activation. J Vest Res 5:137–145 transport and sensitivity. Brain Res 243:215-224 Li YQ, Takada M, Mizuno N (1993) The sites of origin of serotoninergic afferent fibers in the trigeminal motor, facial and hypoglossal nuclei in the rat. Neurosci Res 17:307-313 中国科技论文在线 http://www.paper.edu.cn

521 Warr WB (1975) Olivocochlear and vestibular efferent neurons of Wiklund L, Leger L, Persson M (1981) Monoamine cell distribution the feline brain stem: their location, morphology and number in the cat brain stem. A fluorescence histochemical study with determined by retrograde axonal transport and acetylcholines- quantification of indolaminergic and cell terase histochemistry. J Comp Neurol 161:159–181 groups. J Comp Neurol 203:613–647 Warr WB (1992) Organization of olivocochlear efferent systems in Willard FH, Ho RH, Martin GF (1984) The neuronal types and the mammals. In: Webster DB, Popper AN, Fay RR (eds) The distribution of 5-hydroxytryptamine and enkephalin-like im- mammalian auditory pathway: neuroanatomy. Springer, New munoreactive fibers in the of the North York, pp 410–448 American opossum. Brain Res Bull 12:253–266 Warr WB, Beck-Boche JE, Ye Y, Kim DO (2002) Organization of Yates BJ, Goto T, Bolton PS (1992) Responses of neurons in the olivocochlear neurons in the cat studied with the retrograde caudal medullary raphe nuclei of the cat to stimulation of the tracer cholera toxin-B. J Assoc Res Otolaryngol 3:457–478 vestibular nerve. Exp Brain Res 89:323–332 Warren EH, Liberman MC (1989) Effects of contralateral sound of Yates BJ, Goto T, Kerman I, Bolton PS (1993) Responses of caudal auditory-nerve responses. I. Contributions of cochlear efferents. medullary raphe nuclei neurons to natural vestibular stimula- Hearing Res 37:89–104 tion. J Neurophysiol 70:938–946 Waterhouse BD, Border B, Wahl L (1993) Topographic organization Ye Y, Kim DO (2001) Connections between the dorsal raphe of rat locus coeruleus and dorsal raphe nuclei: distribution of nucleus and a hindbrain region consisting of the cochlear cells projecting to visual system structure. J Comp Neurol nucleus and neighboring structures. Acta Otolaryngol 121:284– 336:345–361 288 White SR, Fung SJ (1989) Serotonin depolarizes cat spinal Ye Y, Machado DG, Kim DO (2000) Projection of the marginal shell motoneurons in situ and decreases motoneuron after-hyperpo- of the anteroventral cochlear nucleus to olivocochlear neurons larizing potentials. Brain Res 502:205–213 in the cat. J Comp Neurol 420:127-138