Substance-P pathways in rat brain.

The characterization and subsequent availability of synthetic substance P has renewed interest in the role of this undecapeptide in central-nervous-system function (Mroz & Leeman, 1974). Biochemical and electrophysiological studies have demonstrated that substance P may function as an excitatory transmitter in primary sensory neurons terminating in the spinal cord (Otsuka & Konishi, 1975). In the brain, however, the available evidence is more difficult to interpret. Using a sensitive radioimmunoassay (Kanazawa & Jessell, 1976), we have previously studied the regional distribution of substance P in the rat brain. Particularly high concentrations of substance P were found in the substantia nigra and interpeduncular nucleus. The interpeduncular nucleus receives afferents from the habenula nuclei (Lenn, 1976), which appear to be primarily cholinergic (Kataska et al., 1973). Substance-Pcontaining cell bodies have been demonstrated immunohistochemically in the habenula nucleus (Hokfelt et al., 1975) and, in addition, lesions of the habenula appear to decrease substance-P concentrations in the interpeduncular nucleus (Mroz et al., 1976). The origin of substance P in the substantia nigra is less clear. The substantia nigra receives a major afferent input from the globus pallidum and corpus striatum (Grofova & Rinvik, 1970) and this pathway is thought to contain y-aminobutyrate (Fonnum et al., 1974). Partial hemisection at the level of the mammillary bodies depletes nigral substance P, suggesting a substance-P pathway originating in either the caudate nucleus or overlying cerebral cortex and passing through the internal capsule (Mroz et al., 1976). The present study was performed to determine more precisely the origin of substance P fibres projecting to the substantia nigra and interpeduncular nucleus. Stereotactic lesions were placed unilaterally in either the corpus striatum or globus pallidum of male 300g Wistar rats. Lesions were made by using a Radionics radiofrequency lesion generator with a tip temperature of 55-56°C. A second series of rats received lesions ablating the medial and lateral habenula bilaterally. Then 7 days after lesioning, the rats were decapitated, the brains removed and the brain stems sliced into 300pm coronal sections with a McIlwain tissue chopper in a cold-room at 4°C. The extent of the lesion was verified histologically after sectioning the brain. Nigral or interpeduncular tissue from two consecutive sections was transferred, as separate samples, to small glass tubes for measurement of substance P, glutamate decarboxylase (EC 4.1.1.15) or choline acetyltransferase (EC 2.3.1.6). Substance P was measured by the extraction procedure and radioimmunoassay described previously (Kanazawa & Jessell, 1976). Glutamate decarboxylase and choline acetyltransferase were assayed essentially as described by Fonnum et al. (1970). After habenula lesions there was a large fall in both the concentration of substance P and choline acetyltransferase activity in the interpeduncular nucleus (see Table 1). A loss of up to 90% of the control choline acetyltransferase activity was found in the interpeduncular nucleus. The decrease in the concentration of substance P paralleled the fall in choline acetyltransferase activity, but was less marked. Approx. 40% of control concentrations of substance P remained after lesions removing 90% of choline acetyltransferase activity. This indicates that there may be a population of substance-Pantaining cells or fibres in the interpeduncular nucleus not directly connected to the habenula nuclei. Parallel immunohistochemical studies also demonstrated a marked loss of substance-P-like immunofluorescence in the interpeduncular nucleus after habenula lesions. It appears, therefore, that the habenula-interpeduncular tract is not solely cholinergic but includes a substance-P pathway also. Biochemical determination of substance P after large pallidal or striatal lesions (see Table 1) suggests that substance-P terminals in the substantia nigra originate in these