Robert J. Dado

The cells of origin of the spinothalamic tract of the rat: a quantitative reexamination

We quantitatively reinvestigated the cells of origin of the spinothalamic tract (STT) of the rat. Injections of Fluoro-Gold that filled the thalamus on one side labeled large numbers of neurons throughout the length of the spinal cord. In 3 cases, we estimated the total number of STT neurons by counting labeled neurons in 18 of the 34 total segments, applying correction factors to these counts, and estimating the numbers of labeled neurons in the 16 remaining unexamined segments. The accuracy of these estimates was tested in two animals in which labeled neurons were counted in all 34 spinal segments. In both cases, the estimated totals of STT neurons differed from the counted totals by less than 5%. In the most effective case, we estimated that more than 9500 STT neurons were labeled. This study indicates that the number of STT neurons in rats is larger than previously reported and suggests that the STT may play an important role in nociception in rats, as it does in primates including humans.

Direct spinal pathways to the limbic system for nociceptive information

The hypothalamus is believed to play important roles in several aspects of nociception. Previously, nociceptive information was thought to reach hypothalamic neurons through indirect, multisynaptic pathways. However, we have found that thousands of neurons throughout the length of the spinal cord in rats send axons directly into the hypothalamus, and many of these axons carry nociceptive information. The axons often follow a complex course, ascending through the contralateral spinal cord, brainstem, thalamus and hypothalamus. They then cross the midline and enter the ipsilateral hypothalamus, turn posteriorly, and continue into the ipsilateral thalamus. These axons might provide nociceptive information to a variety of nuclei in the thalamus and hypothalamus bilaterally.

Evidence that Fluoro-Gold can be transported avidly through fibers of passage.

Small iontophoretic injections of the retrograde tracer Fluoro-Gold were restricted to the dorsal columns in the cervical enlargement of 6 rats. Large numbers of neurons were labeled in the lumbosacral dorsal horn in each rat. In the most effective case, more than 1800 neurons were labeled in alternate sections through nine examined segments. Many neurons were also labeled in lumbosacral dorsal root ganglia of all cases. This study, in contrast to previous reports, indicates that Fluoro-Gold can be transported avidly by axons passing through, but not terminating in, injection sites.

Afferent input to nucleus submedius in rats: retrograde labeling of neurons in the spinal cord and caudal medulla

In cats, spinal and medullary input to the thalamic nucleus submedius (Sm) arises almost exclusively from neurons in the marginal zone. As a result, it has been proposed that Sm may be specifically involved in nociception. In the present study, we determined the locations of neurons in the spinal cord and caudal medulla that project to Sm in rats. Iontophoretic injections of Fluoro-Gold or pressure injections of Fast blue were made into Sm. In each of the 6 rats that received small injections of Fluoro-Gold into Sm, only a small number (mean = 90) of retrogradely labeled neurons were found throughout the 18 segments of the spinal cord examined. Surprisingly, almost no labeled neurons (less than 1%) were counted in the marginal zone of the spinal cord. The majority were located in the deep dorsal horn and intermediate zone/ventral horn. In contrast, many neurons were labeled in the marginal zone of nucleus caudalis. Injections of Fluoro-Gold into any of a number of nuclei near Sm also labeled only a small number of neurons in the spinal cord and almost no neurons in the marginal zone. Using identical injection parameters, we injected Fluoro-Gold into the ventrobasal complex or posterior thalamic group. Hundreds of neurons in the spinal cord, including many in the marginal zone, were labeled following these injections. These results indicate that the techniques used to inject Fluoro-Gold into Sm were capable of labeling many projection neurons, including those in the marginal zone. Larger pressure injections of Fast blue were also made into Sm of 3 rats. The distribution of labeled neurons in nucleus caudalis and the spinal cord was similar to that following iontophoretic injections of Fluoro-Gold. Again, few marginal zone neurons were labeled in the spinal cord in any of these rats. Therefore, our results indicate that few spinothalamic tract neurons appear to project to Sm or any of several adjacent nuclei, and virtually no marginal zone neurons in the spinal cord project to these areas.

Distribution of neuropeptide receptors: New views of peptidergic neurotransmission made possible by antibodies to opioid receptors

The cloning of receptors for neuropeptides made possible studies that identified the neurons that utilize these receptors. In situ hybridization can detect transcripts that encode receptors and thereby identify the cells responsible for their expression, whereas immunocytochemistry enables one to determine the region of the plasma membrane where the receptor is located. We produced antibodies to portions of the predicted amino acid sequences of delta, mu, and kappa opioid receptors and used them in combination with antibodies to a variety of neurotransmitters in multicolor immunofluorescence studies visualized by confocal microscopy. Several findings are notable: First, the cloned delta opioid receptor appears to be distributed primarily in axons, and therefore most likely functions in a presynaptic manner. Second, the cloned mu and kappa opioid receptors are found associated with neuronal plasma membranes of dendrites and cell bodies and therefore most likely function in a postsynaptic manner. However, in certain, discrete populations of neurons, mu and kappa opioid receptors appear to be distributed in axons. Third, enkephalin-containing terminals are often found in close proximity (although not necessarily synaptically linked) to membranes containing either the delta or mu opioid receptors, whereas dynorphin-containing terminals are often found in proximity to kappa opioid receptors. Finally, a substantial mismatch between opioid receptors and their endogenous ligands was observed in some brain regions. However, this mismatch was characterized by complementary zones of receptor and ligand, suggesting underlying principles of organization that underlie long-distance, nonsynaptic neurotransmission.