Physiological identification of pontomedullary serotonergic neurons in the rat

Spinal serotonin is derived entirely from bulbar sources and plays an important role in spinal modulatory processes, including pain modulation. Establishing the electrophysiological properties of SEROTONERGIC bulbospinal neurons in the pontomedullary raphe and reticular formation is critical to understanding the physiological role of serotonin in the spinal cord. Neurons were characterized by their responses to noxious stimulation and their background discharge pattern in the lightly anesthetized rat. Characterized cells were intracellularly labeled with Neurobiotin, which was visualized with a Texas Red fluorophore. Sections containing the labeled cells were processed for serotonin immunocytochemistry with the use of a Bodipy fluorophore. Forty-seven intracellularly labeled cells were tested for serotonin immunoreactivity. The labeled neurons were located in raphe magnus, the nucleus reticularis magnocellularis, and the adjacent reticular and raphe nuclei at levels from the inferior olivary complex to the superior olivary complex. SEROTONERGIC cells were located in the raphe nuclei, in nucleus reticularis magnocellularis pars alpha, and in nucleus reticularis magnocellularis pars beta or nucleus reticularis gigantocellularis. Thirteen intracellularly labeled cells contained serotonin immunoreactivity. The background discharge rate of SEROTONERGIC cells average 1.8 Hz (range: 0.5-3.1 Hz). Discharge was steady and without sustained pauses or bursts in firing. Most serotonin-immunoreactive cells were unaffected or slightly excited by pinch and were unaffected by noxious heat. Three SEROTONERGIC cells were weakly excited by both noxious pinch and heat, whereas two SEROTONERGIC cells were briefly inhibited by these stimuli. Cells that lacked serotonin immunoreactivity were heterogeneous and included ON, OFF, and NEUTRAL cells. Nonserotonergic cells differed from SEROTONERGIC cells in having an irregular discharge pattern and/or a high mean discharge rate. A linear discriminant function, employing background discharge characteristics as independent variables, was calculated that successfully classified 13 of 13 SEROTONERGIC and 32 of 33 nonserotonergic neurons. The probability of misclassification with the use of this discriminant function was estimated to be < 10%. Employing the discriminant function on a test group of cells whose immunochemical content was unknown revealed a population of SEROTONERGIC-LIKE cells that resembled the labeled SEROTONERGIC cells in background discharge pattern, response to noxious stimulation, and nuclear location. The discharge of pontomedullary SEROTONERGIC neurons is slow and steady, suggesting that these neurons may have a role in the tonic, rather than phasic, modulation of spinal processes.