Substance P and Somatostatin-Like Immunoreactivities in Synaptic Glomeruli of the Rat Substantia Gelatinosa, as Revealed by Bi-Specific Monoclonal Antibodies

Abstract

Three distinct types of synaptic glomeruli have been described by one of us in the superficial dorsal horn of rat spinal cord (1,2). These types can be distinguished by the morphology of the central bouton, types of peripheral profiles and distribution in the dorsal horn (1,2). The type I central bouton (C_I) is small, dark and highly scalloped, and prevails in the middle third of lamina II. It is capsaicin sensitive and so is probably of unmyelinated, sensory origin (3). The type IIa glomeruli can be distinguished from type IIb by the presence in the latter of neurofilament bundles (2); both have central endings (C_IIa and C_IIb), that come from myelinated sensory afferents (3), which are lighter, larger and have a smoother surface than the C_I (1,2). In a recent study (4), it was shown that 80% of C_I have FRAP (fluoride resistant acid phosphatase) activity, but no information was available concerning the neurochemical nature of the remaining 20% and of the C_IIa and C_IIb. Based on the above, we decided to study the peptide immunoreactivity in synaptic glomeruli using the bi-specific monoclonal antibodies P₄C₁ (antisubstance P/anti-HRP) (5) and Y₄C₇ (anti-somatostatin/anti-HRP) (6). These antibodies allow a drastic simplification of current immunocytochemical procedures. The incubation with antibodies is reduced to one step. The suppression of link antibodies and PAP complexes results in an intense immunostaining, an increased signal to noise ratio and an excellent penetration of antibodies in the absence of detergents (5). Thus, the ultrastructural preservation is significantly improved. This approach facilitated the correlation of peptide immunoreactive sites with ultrastructural details which define the classification of synaptic glomeruli in the substantia gelatinosa. The precise laminar localization of the FRAP reactive band and somatostatin and substance P-like immunoreactivities was also studied in the superficial dorsal horn of the rat spinal cord.