Auditory afferents terminating as "large myelinated club endings" on goldfish Mauthner cells are identifiable "mixed" (electrical and chemical) synaptic terminals that offer the unique opportunity to correlate physiological properties with biochemical composition and specific ultrastructural features of individual synapses. By combining confocal microscopy and freeze-fracture replica immunogold labeling (FRIL), we demonstrate that gap junctions at these synapses contain connexin35 (Cx35). This connexin is the fish ortholog of the neuron-specific human and mouse connexin36 that is reported to be widely distributed in mammalian brain and to be responsible for electrical coupling between many types of neurons. Similarly, connexin35 was found at gap junctions between neurons in other brain regions, suggesting that connexin35-mediated electrical transmission is common in goldfish brain. Conductance of gap junction channels at large myelinated club endings is known to be dynamically modulated by the activity of their colocalized glutamatergic synapses. We show evidence by confocal microscopy for the presence of the NR1 subunit of the NMDA glutamate receptor subtype, proposed to be a key regulatory element, at these large endings. Furthermore, we also show evidence by FRIL double-immunogold labeling that the NR1 subunit of the NMDA glutamate receptor is present at postsynaptic densities closely associated with gap junction plaques containing Cx35 at mixed synapses across the goldfish hindbrain. Given the widespread distribution of electrical synapses and glutamate receptors, our results suggest that the plastic properties observed at these identifiable junctions may apply to other electrical synapses, including those in mammalian brain.
The numbers of myelinated and unmyelinated fibers were counted in dorsal roots of adult rats treated neonatally with capsaicin in doses ranging from 5 to 100 mg/kg. Substance P and somatostatin levels in the spinal cord, dorsal roots, and sensory ganglia also were determined in control and treated animals. Capsaicin administration lead to the loss of both small myelinated and unmyelinated fibers from dorsal roots. However, whereas a near total loss, up to 94%, of unmyelinated fibers was achieved after high doses of capsaicin, the reduction of myelinated fibers, even of the smallest caliber, did not exceed 40%. The degree of fiber loss showed a clear dose dependency, with little detectable damage to myelinated fibers at doses of less than 50 mg/kg and with an ED50 for damage to unmyelinated fibers of 5 to 10 mg/kg. In all of the structures examined, particularly the dorsal roots, a roughly parallel decrease of substance P and somatostatin was found with capsaicin dose. The depletions of spinal cord substance P (55%) and somatostatin (20%) produced by neonatal capsaicin treatment were similar to those produced by dorsal rhizotomy. Capsaicin does not appear to be specific for primary afferents containing either substance P or somatostatin.
The recently cloned and characterized hyaluronan (HA) receptor RHAMM (receptor for HA-mediated motility) has been shown to play a critical role in mechanisms underlying the motile capacity of a variety of peripheral cell types. Similarities in molecular processes that govern cell locomotion and growth cone migration prompted us to investigate whether RHAMM also contributes to neurite migration in vitro. In immunohistochemical studies of PC12 cells, NG108–15 cells and a neuroblastoma/spinal cord neuronal hybrid cell line (NSC-34 cells) as well as rat and human primary neurons, a punctiform RHAMM labeling pattern was detected in cell bodies, along processes, and at growth cones. By Western blot analysis, the cells lines expressed major RHAMM forms with apparent MW of 60, 75, and 116 kDa. Treatment of NG108–15 cells with dibutyryl-cAMP led to a clear increase in immunolabeling for RHAMM and enhanced expression of the 60 and 75 kDa forms. A polyclonal anti-RHAMM antibody that interferes with HA/RHAMM interaction significantly reduced neurite migration of each cell type examined, while another directed against a RHAMM repeat sequence thought to promote RHAMM receptor aggregation significantly stimulated neurite migration of NSC-34 and rat primary neurons. Different monoclonal anti- RHAMM antibodies had differential inhibitory actions on neurite movement. Low concentrations (ng/ml) of a peptide corresponding to an HA binding domain within RHAMM inhibited neurite migration. These results are the first to implicate RHAMM in the mediation of neurite motility and migration and to point to the potential importance of HA in this process.
The activity of adenosine deaminase (ADA) was measured in 62 discrete regions of the CNS, and in some autonomic and sensory ganglia, peripheral nerves, and peripheral tissues of the rat using an automated high-pressure liquid chromatography (HPLC) method. The formation of inosine and hypoxanthine as a measure of ADA activity in homogenates of brain was optimal at pH 7.0, linear for up to 60 min at 37 degrees C using 500 microM adenosine as substrate, and linear with protein concentrations ranging from 0.05 to 0.8 mg. The Km and Vmax values for ADA activity in homogenates of whole brain were 47 microM and 107 nmol/mg protein/30 min, respectively. Among the CNS regions examined, the highest activity was found in posterior hypothalamic magnocellular nuclei and the lowest in hippocampus. In general, spinal cord contained relatively low levels of ADA activity, with that in dorsal cord approximately 40% higher than ventral cord. In the periphery, parasympathetic ganglia contained higher levels of ADA than sensory ganglia and brain. Most peripheral tissues--including adrenal gland, lung, liver, and anterior and posterior pituitary--exhibited activity comparable to levels in the posterior hypothalamus. ADA activity in thymus was about 10 times higher than any other tissue examined. The uneven distribution of ADA activity in the rat CNS corresponds well with the immunohistochemical localization of this enzyme in discrete neural systems of this species. Structures that contain high ADA activity exhibit intense ADA immunostaining of neuronal perikarya and/or fibers.(ABSTRACT TRUNCATED AT 250 WORDS)
Immunohistochemical analysis of adenosine deaminase in rat brain revealed an extensive plexus of adenosine deaminase-containing neurons in the basal hypothalamus. These neurons converged on and were most numerous in three major centers, namely, the tuberal, caudal, and postmammillary caudal magnocellular nuclei. Most other brain regions were devoid of cells containing adenosine deaminase. Some adenosine deaminase-containing neurons were retrogradely labeled with the fluorescent dye fast blue when the dye was injected into the frontal cortex and striatum. Specific populations of neurons having high levels of adenosine deaminase may release adenosine as a neurotransmitter.
The mode of termination of primary afferent fibres within the superficial dorsal horn of the spinal cord was studied with anterograde tracing methods in normal rats and in animals which had been treated with capsaicin (50 mgkg. s.c.) shortly after birth. In normal animals following injections of horseradish peroxidase (HRP) into lumbar dorsal root ganglia L4, L5, L6, a Golgi-like filling of primary afferent fibres was seen consistently within laminae I, 11, and I11 of the dorsal horn. Several types of laminarelated arborisations were observed, depending upon the survival time after HRP injection. At the earliest survival time (1-2 days) a punctate granular labelling was found in laminae I and I1 inner (I&). At 3 days survival a Golgilike labelling of primary afferent axons occurred and three tiers of arborisation were seen within laminae I, I1 outer (II,), and I1 inner (IIi), respectively.By 7 days after injection this pattern was considerably reduced and there was coarser fibre labelling within laminae I, II,, and 111.When these tracing experiments were repeated in capsaicin-treated rats in which up to 90% of unmyelinated fibres had been destroyed, evidence for a considerable loss of some inputs and rearrangement of the remaining fibres was found. At 3-5 days survival the axonal labelling within laminae I and IIi was severely depleted yet some input, particularly to lamina II,, was present. At 7 days survival deep fibre labelling within lamina 111 now extended well into lamina 11.Mapping of the primary afferent input following 3H-proline injections into the dorsal root ganglion and subsequent autoradiographic processing confirmed that primary afferents terminated throughout the dorsal horn but most heavily in two bands within laminae I and 111. These two dense layers of termination were not present in capsaicin-treated animals, suggesting that they corresponded to areas of unmyelinated primary afferent fibre input.These data, taken together with histochemical observations on the distribution of substance P and fluoride-resistant acid phosphatase (FRAP)-containing primary afferents, suggests that the earliest-labelled fibre systems (3 days) were unmyelinated and replaced at 7 days by predominantly A6 myelinated fibres. It is further suggested that of the three tiers of C fibre terminations seen at 3 days, the most superficial corresponds in part to those primary afferents that can be stained for substance P within lamina I while the most ventral tier within lamina IIi are those primary afferents containing FRAP. The presence of the two temporally distinct fibre systems also suggests that laminae I and 11, receive both C and A6 primary afferent input while lamina 11, receives only C fibre input.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.