The Caudal Raphe Nuclei

Raphe nuclei are cellular groups present in the midline of the brain stem which extend rostrocaudally from the level of the interpeduncular nucleus in the midbrain to the level of the pyramidal decussation in the medulla. These nuclei are either separated from the surrounding areas of the brain stem tegmentum, or tend to merge into the neighbouring areas of the brain stem reticular formation (rats, Steinbusch and Nieuwenhuys, 1983; rabbits, Felten and Cummings, 1979; kittens, Taber et al., 1960). The terminology of caudal raphe nuclei and morphology of neurones of caudal raphe nuclei are briefly described below. The caudal raphe nuclei are the raphe pallidus (RP), obscurus (RO) and the most caudal part of raphe magnus (RM).

Raphe pallidus (RP) is a rod-like nucleus which is situated close to the ventral surface o f the medulla and located between the two pyramids. The nucleus is ventral to, but separated from the RO by fibres of the medial lemniscus and the olivocerebellar tract, as well as by the inferior olivary nucleus. The RP extends rostrally to the caudal levels of the RM which corresponds to a level through the middle of the facial nucleus and caudally to the dorsomedial edges or the pyramidal decussation. Its ventral border touches the basal surface of the brain. The RP is relatively compact. Taber et al. (1960) reported that the caudal RP can be subdivided into a dorsal and a ventral group in the cat and three types of cells may be distinguished in the RP using Nissl stained sections. In the rat, two cell types were found using cresyl-violet stained sections: medium-sized, round/oval neurones with a diameter less than 18 /xm, and elongated/fusiform, large cells with a diameter more than 36 /xm. The predominant portion of neurones are o f medium size and are found in the ventral part of RP (Steinbusch and Nieuwenhuys, 1983; Tork, 1985). The direction of their dendrites varies but for cells situated in the dorsal part of the RP they project dorsally or ventrolaterally.

The nucleus raphe obscurus (RO) is a relatively narrow mid-sagittal cell group which is arranged in two paramedian lines in the middle of the medulla, dorsal to the RP.

Caudally the nucleus extends as far as the medulla itself. It begins slightly above the caudal pole of the inferior olive and its caudal-most cells are intermingled between fibres of pyramidal decussation. Rostrally the nucleus can be followed to approximately the level of the caudal pole of the facial nucleus, overlaps with the caudal part of RM, but its organization is different to that of the RM. The cells in the RO form two distinct parallel and vertical laminae adjacent to the midline. In the rat, RO contains two types of neurones, small spherical neurones with a diameter of approximately 17 /xm and large bipolar or tripolar neurones with a diameter ranging from 28 to 36 /xm. The neurones are distributed evenly within the nucleus and usually have three to four dendrites which are directed dorsalaterally (Steinbusch and Nieuwenhuys, 1983).

The region of the brainstem containing the raphe nuclei has been shown to project to the somatosensory regions of dorsal horn, the motoneurones of the ventral horn and the IML, which implicates their involvement in the central control of various functions (Basbaum et al., 1978). In the last decade the involvement of the raphe nuclei in autonomic regulation has received considerable attention.

In the early studies, of the spinal projections from the raphe regions were reported following spinal degeneration (Torvik and Brodai 1957; Brodai et al., 1960). Using histofluorescence techniques combined with lesions made in the spinal cord, Dalhstrom and Fuxe (1964) found that the IML received a dense input from a bulbospinal serotonergic-containing neurons of the brain stem. Spinal projections of raphe nuclei were further confirmed by retrograde labelling, injecting horseradish peroxidase (HRP) into various regions of the spinal cord in the rats (Basbaum and Fields, 1979; Loewy and McKellar, 1981), cats (Kuypers and Maisky, 1975; Martin at al., 1978), rabbit (Haselton et al., 1988) pigeons (Cabot at ah, 1982) and the North American Opossum (Crutcher et al., 1978). Cabot, Wild and Cohen (1979) used retrograde transport of HRP applied into the spinal cord, anterograde transport-pH]- proline applied into raphe nuclei and degeneration studies in the pigeon, and confirmed the raphe-spinal pathway.

Direct evidence of neurones in caudal raphe nuclei projecting to the IML was obtained by the anterograde autoradiographic technique in the rat. Following injection of the [^H] amino acids tracers ([^HJproline, [^Hjleucine, and [^Hjlysine) into the raphe pallidus and raphe obscurus, it was found that both raphe pallidus and raphe obscurus projected to the IML (Loewy, 1981). Some other studies using anterograde labelling (Basbaum et al., 1978; Holstege and Kuypers, 1982; Martin et al., 1981) and retrograde labelling techniques (Bowker et al., 1981; Martin et al., 1981; Skagerberg and Bjorklund, 1985) revealed more detail of heterogeneous spinal terminations of raphe nuclei. The more caudally located RP and RO neurones project mainly to the motoneurones of ventral horn and the IML, while the RM project predominantly to the dorsal horn.

The transneuronal virus (herpes virus suis, pseudorabies) labelling technique has also shown that neurones in caudal raphe nuclei probably regulate SPNs projecting to various sympathetic ganglia and the adrenal gland in rats (Strack et al., 1989a,b).

Direct evidence of synaptic contacts between raphe neurones and identified SPNs was obtained recently. Phaseolus vulgaris leucoagglutinin (PHA-L), an anterograde tracer was injected into the region of the raphe nuclei of the rat. Synaptic contacts were found between PHA-L anterograde-labelled terminals and preganglionic neurons retrogradely labelled from the adrenal medulla (Zagon et al., 1989). The descending axons gave rise to a widespread distribution of fibres in the grey matter of both the dorsal and ventral horns the IML, IC and CA and occasionally to the contralateral side (Bacon et al., 1990).

1.3.3b Physiological studies

Inhibitory effect o f the caudal raphe area on sympathetic activity

The medullary raphe used to be referred to as a depressor area (Wang and Ranson, 1939; Alexander, 1946; Gootman and Cohen, 1970). Focal stimulation in the raphe area decreased blood pressure, heart rate and sympathetic nerve discharge (Henry and Calaresu, 1974; Coote and Macleod, 1974; Gilbey et al., 1981). Raphe lesions

produced a marked enhancement of the conditioned cardioacceleration response in pigeons (Cabot et al., 1979) and significant increases in spontaneous sympathetic activity in cats (McCall and Harris, 1987).

The inhibitory effect of raphe stimulation on either preganglionic or postganglionic sympathetic nerve activity has been observed in many species: cat, rat and pigeons (Coote and Macleod, 1975; Yusof and Coote, 1988; Cabot et al., 1979). Evidence of an inhibitory effect on SPNs recorded extracellularly was found following stimulation of raphe nuclei in the pigeon (Cabot et al., 1979).

Evidence o f heterogenous effects due to stimulating within the raphe nuclei

Although raphe spinal inhibition has been well documented, results from other experiments demonstrated that there was also a sympatho-excitatory response to raphe stimulation.

Stimulation of caudal raphe nuclei produced both pressor and depressor responses (cat, McCall, 1984). Further studies on the precise anatomical substrates in the caudal raphe area in the cat have shown that electrical stimulation of different areas of raphe could evoke either depressor or pressor responses (Adair et al., 1977). Depressor responses were obtained only after stimulating the anterior region of raphe obscurus or posterior region of raphe magnus and while the posterior region of raphe obscurus produced pressor responses. The nucleus raphe pallidus appears to be a heterogeneous region producing both pressor and depressor responses to stimulation.

Single shock stimulation in pressor sites produced an excitatory evoked potential in the inferior cardiac sympathetic nerve discharge was reported by McCall (1984). On the unanaesthetized decerebrate cat, Futuro-Neto and Coote (1982b) observed various responses of sympathetic nerves innervating heart, renal and skeletal muscle induced by electrical stimulation of the raphe obscurus; an increase of the activity of sympathetic nerve to muscle whereas a decrease o f kidney which similar to a pattern of sympathetic activity during desynchronized sleep-like periods (Futuro-Neto and

Coote, 1982a; Coote, 1982).

In the rat other experiments reported that electrical stimulation of the raphe obscurus yielded either inhibition or excitation of renal and skin sympathetic nerve activity, or yielded diverse effects on these nerves i.e. a decrease in renal sympathetic nerve activity and an increase in muscle sympathetic nerve activity (Yusoff and Coote, 1988). The above studies suggest that the caudal raphe nuclei are heterogeneous in structure and subserve several functions.

To understand the diverse results obtained from various experiments, it should be noted that most of the evidence was obtained by means of electrical stimulation within the brain stem. However, electrical stimulation always has the problem that it cannot be ascertained as to whether or not stimulation is of neurones or fibres of passage. Recent experiments in the rat were conducted by using both electrical and chemical stimulation; glutamate injection into raphe obscurus caused a rise in blood pressure but no bradycardia whereas electrical stimulation caused both a rise in blood pressure and bradycardia (Dreteler et al., 1991). It is likely that the bradycardia caused by electrical stimulation in this experiment was probably due to stimulation of axons of passage.