Effects of MK-771 on the isolated amphibian spinal cord: comparison with thyrotropin-releasing hormone.

Topically applied MK-771 (pyro-2-aminoadipyl-histidyl-thiazolidine-4-carboxamide), a novel thyrotropin-releasing hormone (TRH) analog, was found to be equipotent with TRH in depolarizing the ventral roots of the isolated, hemisected amphibian (Bufo marinus) spinal cord. The 3-methyl-histidyl analog of TRH was approximately 10 times more potent than MK-771 and TRH. MK-771 is known to be equiactive with TRH in their actions on the pituitary gland. Taken together these findings suggest that the previously observed enhanced potency of systemically administered MK-771 over TRH in in vivo central nervous system (CNS) test paradigms is not likely to be due to a difference in the agonist requirements of CNS as compared with pituitary receptors for TRH.     

Effect of serotonin on isolated cells with the various functionality from the lamprey spinal cord

The differential actions of 5-hydroxytryptamine (5-HT) (100 microM) were investigated on isolated motoneurons, interneurons, and primary sensory neurons from the lamprey spinal cord using patch-clamp techniques. Application of 5-HT did not evoke membrane currents in any of the spinal neurons tested (n = 62). However, in most motoneurons and interneurons (15 of 18), 5-HT produced a small depolarization (2-6 mV), which was not accompanied by a change in input resistance. In the remaining motoneurons and interneurons (3 of 18), 5-HT induced a large depolarization (up to 10-20 mV) and a decrease in input resistance of 20-60%. In most sensory neurons (dorsal sensory cells, DSCs), 5-HT evoked a short-lasting, low-amplitude depolarization, followed by a long-lasting hyperpolarization of 2-7 mV. The DSCs showed no significant change in input resistance to 5-HT application (n = 8). Spike afterpolarization were also differentially modulated by 5-HT. In motoneurons and interneurons, 5-HT decreased the amplitude of the afterhyperpolarization following the action potential while increasing the amplitude of the after depolarization. In the DSCs, no significant effect of 5-HT on spike afterpolarization was observed. 5-HT differentially modulated the current induced by application of N-methyl-D-aspartate (NMDA). In motoneurons and interneurons, 5-HT enhanced NMDA-evoked current, while in DSCs, 5-HT decreased this current. These results demonstrate that 5-HT differentially modulates the activity of functionally different groups of spinal neurons. In motoneurons and interneurons, 5-HT enhances excitation by inducing depolarization and decreasing the afterhyperpolatization, while NMDA currents are enhanced. These effects facilitate the appearance of rhythmic discharges in these cells in the presence of NMDA. In primary dorsal sensory cells, 5-HT enhances inhibition by hyperpolarizing the cells and depressing NMDA currents. These differential effects are presumably mediated by different types of 5-HT receptors on these classes of spinal neurons.     

The neuroanatomy of an amphibian embryo spinal cord

Horseradish peroxidase has been used to stain spinal cord neurons in late embryos of the clawed toad (Xenopus laevis). It has shown clearly the soma, dendrites and axonal projections of spinal sensory, motor and interneurons. On the basis of light microscopy we describe nine differentiated spinal cord neuron classes. These include the Rohon-Beard cells and extramedullary cells which are both primary sensory neurons, one class of motoneurons that innervate the segmental myotomes, two classes of interneurons with decussating axons, three classes of interneurons with ipsilateral axons and a previously undescribed class of ciliated ependymal cells with axons projecting ipsilaterally to the brain. We believe that all differentiated neuron classes are described and that this anatomical account is the most complete for any vertebrate spinal cord.     

Identification of neuropeptide FF-related peptides in rodent spinal cord.

Peptides which should be generated from the neuropeptide FF (NPFF) precursor were identified in mouse and rat spinal cord, by using reverse phase high pressure liquid chromatography with radioimmunoassay and electrospray mass spectrometry detection. In both species, two octapeptides, NPFF (Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide) and NPSF (Ser-Leu-Ala-Ala-Pro-Gln-Arg-Phe-amide) were identified but a longer peptide NPA-NPFF (Asn-Pro-Ala-Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide) was present at the highest concentration in rat spinal cord. In mouse, the homologous peptide, SPA-NPFF (Ser-Pro-Ala-Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide) was not detected. Both peptides NPFF and NPSF reverse morphine-induced analgesia in the tail flick test. Our data reveal species differences in the maturation of NPFF precursor.     

The effects of various peptides on the isolated pulmonary artery

Helical strips of pulmonary arteries from rabbits were tested for their responses to the following peptides: neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), substance P (SP), cholecystokinin-octapeptide (CCK), somatostatin (SS), bombesin, neurotensin and gastrin. SP, in the absence of active base-tension, and NPY both induced concentration-dependent contractions, while VIP and SP, in the presence of active base-tone, and CCK induced relaxation. The pD2 (-log ED50) was in the order of NPY greater than SP and SP greater than CCK greater than VIP. SS, bombesin, NT and gastrin had no effect. These findings suggest peptidergic involvement in the vessels.     

Effects of barium on isolated frog spinal cord

The effects of Ba2+ were studied in vitro on the isolated frog spinal cord. Ba2+ (25 microM-5 mM) caused a concentration-dependent depolarization of ventral (VR) and dorsal (DR) roots. TTX and Mg2+ substantially reduced the depolarization suggesting that interneuronal effects were involved. Ba2+ (25-500 microM) markedly increased the frequency and duration of spontaneous VR and DR potentials and substantially enhanced the duration (and frequently the amplitude) of VR and DR potentials evoked by DR stimulation. Higher concentrations of Ba2+ (1-5 mM) reduced both spontaneous and evoked potentials. Ba2+ (25-500 microM) enhanced the amount of K+ released by a DR volley and by application of L-glutamate and L-aspartate. The cation reduced VR and DR root depolarizations produced by elevated [K+]0. VR potentials induced by L-glutamate, L-aspartate, GABA and glycine and DR depolarizations caused by GABA were reduced by Ba2+. These results show that Ba2+ has complex actions on reflex transmission, interneuronal activity, the postsynaptic actions of excitatory and inhibitory amino acids and the evoked release of K+.     

Actions of gastrin-releasing peptide and related mammalian and amphibian peptides on ion transport in the porcine proximal jejunum

The 27-amino acid peptide gastrin-releasing peptide (GRP) and the decapeptide neuromedin B (NMB) are structurally related to bombesin (BB) and exist within the mammalian small intestine. We examined the actions of porcine GRP and NMB on ion transport in the porcine proximal jejunum in vitro and compared their activities to those of their respective C-terminal amphibian homologs BB and ranatensin (RT). The 4 peptides transiently increased potential difference and short-circuit current (Isc) in jejunal mucosal sheets after their serosal administration in subnanomolar concentrations with an order of potency: GRP approximately RT greater than or equal to NMB greater than BB. BB and RT were more effective in elevating Isc than GRP and NMB; all peptides had variable effects on tissue conductance. Mucosal Isc responses to GRP (1 nM) were due in part to a stimulation of net Cl- secretion. GRP-induced Isc increases were halved by serosal furosemide (0.3 mM) and reduced by 65% and 90% in tissue bathing solutions lacking Cl- or Cl- and HCO3-, respectively. Tetrodotoxin reduced Isc responses to the peptide by 40%; GRP activity remained unaffected after blockade of gut muscarinic or nicotinic cholinergic receptors by atropine or hexamethonium, respectively. These results suggest that GRP and its natural homologs stimulate active electrogenic Cl- secretion in the porcine jejunum through interactions with GRP receptors located in the intestinal mucosa and submucosa.     

Effect of monoamines on motoneurons of the isolated rat spinal cord

Superfusion of isolated hemisected spinal cords of 9-13-day old rats with noradrenalin (NA) solution depolarized or hyperpolarized the motoneurons depending on the NA concentration. Both effects were the result of the direct action of NA on the motoneurons, for they were given in medium containing an excess of Mg and deficiency of Ca ions.a-Adrenoblockers depressed both the depolarizing and hyperpolarizing effects of NA. The depolarizing effect of dopamine on motoneurons was abolished in medium containing excess of Mg ions. Its direct hyperpolarizing action of motoneurons was suppressed by haloperidol but unchanged by phentolamine. The depolarizing effect of serotonin and its metabolites — mexamine, kynurenine, and 3-hydroxy-anthranilic acid — persisted in the presence of an excess of Mg and deficiency of Ca ions, but it was suppressed by deseryl (methysergide) and the benzyl analog of serotonin. The hyperpolarizing effect of serotonin at high concentrations (10^–4–10^–3 M), revealed in some experiments, was abolished in medium containing excess of magnesium ions in the presence of morphine.A. M. Gorkii Donetsk State Medical Institute. Translated from Neirofiziologiya, Vol. 12, No. 4, pp. 391–396, July–August, 1980.     

Characterization of beta-endorphin peptides in the spinal cord of the rat

The objective of the present studies was to estimate the total content of beta-endorphin-like immunoreactivity (beta-EPLIR) and to characterize the beta-endorphin-like peptides in distinct regions of the spinal cord using gel filtration and reverse phase high performance liquid chromatography. The concentration of beta-EPLIR expressed as pg per mg tissue was similar in the four regions of the spinal cord. Sephadex G-75 chromatography demonstrated the presence, in all four regions of the spinal cord, of beta-endorphin (beta-EP) immunoreactive peptides eluting at the positions of standard beta-EP and beta-lipotropin (beta-LPH) peptides as well as a high molecular weight form eluting prior to beta-LPH. High performance liquid chromatography of the beta-EP-sized peptides indicated some differences in the relative proportions of the various beta-EP-sized peptides among the four regions of the spinal cord, which suggest a different origin of the beta-EP fibers terminating in different regions of the spinal cord as well as different physiological importance of the beta-endorphin peptides in the various spinal cord regions.     

Electrotonic transmission between primary afferent fibers and the motor neurons of the isolated spinal cord of various representative amphibians

Studies have been made of monosynaptic excitatory postsynaptic potentials elicited by stimulation of the posterior root in motoneurones of the isolated spinal cord of the frog Rana ridibunda, toad Bufo bufo, and clawed toad Xenopus laevis. In all the amphibians studied, the early component of monosynaptic EPSP was not blocked in Ca-free medium containing 2 mM Mn2+. It is suggested that electrical coupling in anuran amphibians reflects certain stage of evolution of the synaptic transmission in vertebrates.     

Influence of gastrointestinal peptides on bile acid transport by isolated rat hepatocytes

While numerous effects of gut peptides on gastric, pancreatic, and intestinal secretion have been described, there has been little investigation of the influence of these peptides on hepatic function. In the present studies, effects of vasoactive intestinal peptide (VIP), somatostatin, thyrotropin-releasing hormone (TRH), and bombesin on taurocholate transport by isolated rat hepatocytes have been examined. Somatostatin, TRH, and bombesin in incubation media produced no change from control incubations with regard to either uptake of taurocholate by hepatocytes or efflux of bile acid from preloaded cells. However, incubation of hepatocytes with VIP produced a significant decrease in taurocholate uptake (1.34 +/- 0.13 versus 1.73 +/- 0.16 nmole.min-1.10(6) cells-1, P less than 0.001). Studies with verapamil, a calcium-channel blocking agent, and theophylline, an inhibitor of cAMP catabolism, failed to provide evidence for transmembrane Ca2+ flux or alteration in intracellular levels of cAMP, respectively, as mechanisms for the observed inhibition of hepatocyte taurocholate uptake by VIP. These data, coupled with both clinical and other basic observations, suggest that VIP may play a significant role in the regulation of hepatic bile secretion.     

Formation of spinal cord cicatrix under various experimental conditions

Structure of the cat spinal cord scar has been studied by means of light and electron microscopy after its lateral hemisection, complete dissection and hemisection in combination with autotransplantation of the sympathetic ganglion, which keeps its connection with the sympathetic trunk, into the cut of the spinal cord. Three zones are revealed in the scar: central (connective tissue), intermediate (glio-connective tissue) and peripheral (zone of glio-cystous and reactive changes of the nervous tissue). Peculiarities of intercellular reactions are revealed in the process of formation of various zones in the scar and their dependence on the type of the experiment. In the experiments with autotransplantation of the sympathetic ganglion into the spinal cord, a definite possibility to restrict scarry changes of the spinal cord is demonstrated in connection with improving reinnervation and revascularization of the traumatized segment.     

Pharmacology of amphibian opiate peptides

In 1980 the skin of certain frogs belonging to the genus Phyllomedusinae was found to contain two new peptides that proved to be selective mu-opioid agonists, and named dermorphins. Since 1987 deltorphins, a family of highly selective delta-opioid peptides were identified either by cloning of the cDNA from frog skins or isolation of the peptides. The distinctive feature of opioid peptides is the presence of a naturally occurring D-enantiomer at the second position in their common N-terminal sequence, Tyr-D-Xaa-Phe. The discovery of the amphibian opiate peptides, provided new insights into the functional role of the mu- and delta-opiate systems. It also provided models for novel analgesics with enhanced therapeutic benefits and reduced toxicity.     

Effect of neuroactive peptides on labeled 5-hydroxytryptamine release from rat spinal cord

The effects of neuroactive peptides on the release of 5-HT were studied. The 5-HT released from the spinal cord was significantly increased by somatostatin, substance P and peripheral pain stimulation (tail pinch), but not affected by neurotensin, beta-endorphin and met-enkephalin. The somatostatin-evoked 5-HT release was inhibited by baclofen and met-enkephalin in vivo but not in vitro. The substance P-evoked 5-HT release was strongly inhibited by baclofen, and slightly potentiated by met-enkephalin in vivo but not in vitro. The tail pinch-induced 5-HT release was inhibited by met-enkephalin and baclofen, but potentiated by naloxone. These findings provide further evidence on the important role of neuropeptides and suggest that the descending serotonergic neurones are modulated by neuropeptide interneurones in the spinal cord.     

The interaction of porphyrin precursors with GABA receptors in the isolated frog spinal cord.

Both delta aminolevulinic acid (ALA) and porphobilinogen (PBG) depressed spontaneous activity in the isolated hemisected frog spinal cord. ALA was approximately one fifth as potent as GABA in this respect and had a much slower time course. ALA hyperpolarized motoneurones and this effect was blocked by the GABA antagonist, picrotoxin, but not by the glycine antagonist, strychnine. ALA also depolarized primary afferents and this effect was similar to that of GABA although its action was somewhat reduced by strychnine. These results suggest that ALA activates the GABA receptors on motoneurones and primary afferents. The possible significance of these results to the clinical manifestations of acute attacks of porphyria is discussed.