bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification and characterization of pleural neurons that inhibit tail sensory neurons and motor neurons in Aplysia: correlation with FMRFamide immunoreactivity
Authors: Xu Y , Cleary LJ , Byrne JH
Published: April 2018

Neurons on the rostral edge of the ventral surface of the right pleural ganglion were identified as elements of the circuit mediating the defensive tail withdrawal reflex of Aplysia. These neurons produced IPSPs in tail sensory neurons and were classified into two groups, RPI4 and RPI5, according to their affinity for an antibody directed against FMRFamide. RPI4 was not FMRFamide immunoreactive, and RPI5 was. RPI4 and RPI5 were found to have different electrophysiological profiles. The summated IPSPs in sensory neurons produced by RPI4 developed more rapidly and had a shorter duration than those produced by RPI5. In addition, RPI4 produced IPSPs in the tail motor neurons, whereas RPI5 did not. Both RPI4 and RPI5 received excitatory synaptic inputs from stimulation of the pleural-abdominal connective as well as peripheral nerves P8 and P9, which innervate the tail and posterior part of the animal's body. These inputs were sufficient to elicit spikes. In RPI4, the excitatory synaptic inputs were followed by short and transient hyperpolarization, whereas in RPI5, the excitatory synaptic inputs were followed by slow and long-lasting hyperpolarization. Excitatory inputs elicited in RPI4 by stimulation of peripheral nerves appeared to be mediated, at least in part, by activation of tail sensory neurons. Intracellular stimulation of sensory neurons produced EPSPs in RPI4 that appeared to be monosynaptic. These results suggest that inhibitory interneurons underlying the circuit of the tail withdrawal reflex may play roles in mediating or modulating neuronal responses to tail stimulation. By inhibiting tail sensory and motor neurons, these interneurons may reduce the effectiveness of an animal's response to stimulation of the tail.
Distribution and synaptic localization of immunocytochemically identified NMDA receptor subunit proteins in sensory-motor and visual cortices of monkey and human
Authors: Huntley GW , Vickers JC , Janssen W , Brose N , Heinemann SF , Morrison JH
Published: April 2018

NMDA receptors are composed of multiple receptor subunit proteins, of which NMDAR1 appears to be a critical component for normal receptor function (Nakanishi, 1992). In this study, quantitative immunocytochemical methods were used at the light and electron microscopic levels to localize NMDAR1 subunits in the primary motor (M1) and somatic sensory (S1) cortex of monkeys, and in the primary visual cortices (V1) of monkey and human. Three principal features of NMDAR1 subunit organization were examined in detail in the monkey cortex: (1) the laminar and cellular distribution patterns, relying in part on double-labeling paradigms with the calcium-binding proteins parvalbumin (PV) and calretinin (CR) as markers for discrete subpopulations of GABAergic interneurons; (2) the codistribution of NMDAR1 subunits with non-NMDA ionotropic receptor subunits; (3) a quantitative assessment of the percentages of asymmetrical synapses in layers II/III, IV, and V/VI that were NMDAR1 immunoreactive. In monkey M1, S1, and V1, NMDAR 1 immunoreactivity was present in all layers, localized primarily to large numbers of pyramidal cell somata and proximal apical dendrites, to presumptive spiny stellate cells in layer IV of V1, and to the vast majority (approximately 80–90%) of PV- immunoreactive cells. By contrast, NMDAR1 immunoreactivity was present in only a very small percentage of the CR-immunoreactive cells (approximately 6–9%). Colocalization with non-NMDA receptor subunits showed that all cells (100%) that contained GluR2/3 subunits were also NMDAR1 immunoreactive. In addition, the complete codistribution of GluR5/6/7 subunits with GluR2/3 subunits suggests, indirectly, that all GluR5/6/7-immunoreactive cells are also NMDAR1 immunoreactive. The laminar and cellular distribution patterns of immunostaining in human V1 were very similar to those in monkey V1. Electron microscopy of monkey sections confirmed an extensive dendritic and synaptic localization of NMDAR1 subunits. Labeling of synapses was present on asymmetrical postsynaptic densities associated with both dendritic shafts and spines. In supragranular layers of V1, a greater percentage of asymmetrical synapses were NMDAR1 immunopositive (44%) in comparison to layer IVC beta (34%) or deep layers (19%). In contrast, in area 3b of S1, the percentage of labeled synapses was greatest in layer IV (45%) in comparison to superficial (26%) and deep (37%) layers, while in M1, the percentages of labeled synapses were similar between superficial (46%) and deep (40%) layers. Taken together, these data indicate that NMDAR1-immunoreactive cells in neocortex represent a morphologically, functionally, and neurochemically heterogeneous population.(ABSTRACT TRUNCATED AT 400 WORDS)
Serotonin modulates photic responses in the hamster suprachiasmatic nuclei
Authors: Rea MA , Glass JD , Colwell CS
Published: April 2018

The aim of the present study was to examine the effects of serotonin agonists on three elements of the photic response in the hamster suprachiasmatic nuclei (SCN). Both serotonin and the selective 5-HT1A agonist 8-OH-DPAT inhibited field potentials recorded in the SCN in response to optic nerve stimulation in the hypothalamic slice preparation. The effects of both drugs were dose related over a concentration range of 1–50 microM, and, in both cases, a maximal inhibition of approximately 60% was achieved at a concentration of 25– 50 microM. Systemic administration of 8-OH-DPAT inhibited light- stimulated Fos expression in the SCN. A regionally selective pattern of inhibition was observed, with decreases restricted predominately to the ventral and dorsal borders of the SCN. Finally, systemic administration of 8-OH-DPAT was found to dose-dependently attenuate light-induced phase shifts of the free-running activity rhythm. The effects of 8-OH- DPAT on light-induced phase advances were dose dependent. Injection of 8-OH-DPAT at a dose of 0.5 mg/kg caused 57% inhibition of light-induced phase advances, while a dose of 5 mg/kg inhibited the phase advance by 82%. Injection of 0.05 mg/kg 8-OH-DPAT did not significantly inhibit light-induced phase advances. Injection of 5 mg/kg 8-OH-DPAT alone did not significantly alter the phase of the activity rhythm. Similarly, injection of 5 mg/kg 8-OH-DPAT 30 min prior to light stimulation at CT14 completely inhibited light-induced phase delays, while this dose of the drug did not alter the phase of the activity rhythm when administered alone.(ABSTRACT TRUNCATED AT 250 WORDS)
Injury-induced plasticity of spinal reflex activity: NK1 neurokinin receptor activation and enhanced A- and C-fiber mediated responses in the rat spinal cord in vitro
Authors: Thompson SW , Dray A , Urban L
Published: April 2018

A- and C-fiber evoked ventral root potential (VRP) responses have been examined in isolated spinal cord preparations maintained in vitro that were taken from young rats in which behavioral hyperalgesia (thermal and mechanical) was induced following UV irradiation of one hindpaw. Evoked VRPs were compared with those in naive untreated animals. The duration of both the A- and C-fiber evoked VRP was significantly increased in UV-treated animals. The amplitude of the summated VRP evoked by repeated low-frequency (1.0–5.0 Hz) C-fiber stimulation, a measure of windup, was significantly greater in UV-treated animals. In UV-treated animals, repeated low-frequency (1.0–5.0 Hz) stimulation of A-fiber inputs to the spinal cord also evoked a significant summated VRP, which was not observed in spinal cords from untreated animals. In naive animals the prolonged VRP evoked following single shock C-fiber stimulation was significantly antagonized by the NMDA receptor antagonist D-AP5 and the NK2 receptor antagonist MEN, 10376 but not by the NK1 receptor antagonists CP-96,345 or RP,67580. Summated VRPs evoked by repeated C-fiber stimulation in naive animals were significantly antagonized only by D-AP5. In hyperalgesic animals the prolonged VRP evoked by C-fiber stimulation was significantly reduced by NK1, NK2, and NMDA antagonists. The summated VRP was also significantly reduced by these antagonists. In both untreated and UV- irradiated animals the single shock evoked A-fiber ventral root response was significantly antagonized only by D-AP5. However, the summated VRP evoked by repeated A-fiber stimulation in UV-treated animals was also significantly reduced by NMDA, NK1, and NK2 receptor antagonists. The present study has demonstrated enhanced A- and C-fiber evoked responses in the rat spinal cord in vitro following induction of a peripheral injury by UV irradiation and which was associated with behavioral hyperalgesia to thermal and mechanical stimuli. Under this condition, repetitive stimulation of A-fiber primary afferents was capable of producing an enhancement of spinal excitability similar to that evoked by C-fibers in normal animals. Furthermore, we have observed the expression of an NK1 receptor component to the C-fiber evoked response following the establishment of the peripheral injury. The enhanced ventral root responses and changes in receptor sensitivity may contribute to the phenomenon of central sensitization and may be directly related to the behavioral hyperalgesia observed. Moreover, these findings may be relevant to the mechanisms of enhanced central excitability that occur in clinical conditions of inflammatory hyperalgesia and neuropathic pain.
Postsynaptic modulation of synaptic efficacy at mixed synapses on the Mauthner cell
Authors: Pereda AE , Nairn AC , Wolszon LR , Faber DS
Published: April 2018

Extracellular application of dopamine in the synaptic bed of the lateral dendrite of the goldfish Mauthner (M-) cell enhances both the electrical and chemical components of the mixed excitatory postsynaptic potential (EPSP) evoked by ipsilateral eighth nerve stimulation (Pereda et. al., 1992). We describe here results of experiments designed to determine the locus of action of dopamine and the underlying cellular mechanisms. This amine acts independently on the two modes of transmission, since (1) the percentage increases in the two were not correlated, (2) the time courses of their modifications were independent, and (3) the observed increases in synaptic responses cannot be attributed to a generalized effect on M-cell input conductance, which was increased by dopamine, a change that would rather be expected to shunt the synaptic potentials. Also, dopamine does not produce presynaptic spike broadening and does not modify paired-pulse facilitation, two indications that it acts postsynaptically. The alterations in the mixed EPSP are presumably due to activation of a postsynaptic cAMP-dependent phosphorylation pathway. Specifically, they did not occur if the cAMP-dependent protein kinase inhibitor PKI5–24 was injected intradendritically prior to dopamine application, and they could, on the other hand, be mimicked by injections of the catalytic subunit of the cAMP-dependent protein kinase, PKACAT. In contrast, neither manipulation altered the M-cell input conductance directly or affected the dopamine-induced increase in conductance, suggesting this effect of dopamine is cAMP independent.(ABSTRACT TRUNCATED AT 250 WORDS)
Intraventricular administration of BDNF increases neuropeptide expression in newborn rat brain
Authors: Nawa H , Pelleymounter MA , Carnahan J
Published: April 2018

Brain-derived neurotrophic factor (BDNF) specifically enhances and maintains the expression of neuropeptide Y (NPY) and somatostatin (SOM) in cultured neocortical neurons (Nawa et al., 1993). In this article, we examined its effects in vivo on neuropeptide expression in various brain regions by injecting BDNF into the cerebroventricle of newborn rats. Repeated administration (2x) of BDNF increased contents of NPY- like immunoreactivity (NPY-LI) and substance P (SP)-LI most markedly in the anterior neocortex by 11- and 24-fold, respectively, in comparison to values in the animals receiving control injection. A smaller but significant increase was also observed in immunoreactivity for somatostatin (SOM), enkephalin (ENK), and cholecystokinin (CCK). mRNA for NPY, SP, and SOM was similarly upregulated in the anterior neocortex, suggesting that BDNF enhances peptide synthesis rather than inhibiting peptide release or degradation. Among the brain regions examined, however, peptidergic responses to BDNF were different with respect to their spatial distribution and time course. Induction of SP- LI, NPY-LI, and SOM-LI around the injection site was most pronounced in cortical layers II/III, layers IV-VI, and layer VI, respectively. Peptidergic immunoreactivity was also enhanced in other brain regions ipsilateral to the injection site, for example, NPY-LI in the hippocampus, thalamic nuclei, and striatum, and SOM-LI in the striatum. A single injection of BDNF elevated SP-LI to a plateau level within 12 hr while NPY-LI and SOM-LI reached maximum levels at 48 hr, and then all returned to control levels at 68 hr. In contrast, the same dose of NGF had no influences on the neuropeptide levels at 48 hr. These observations suggest that BDNF regulates the development of neuropeptide expression in the CNS in a plastic manner.
Developmental regulation of the hypothalamic metabotropic glutamate receptor mGluR1
Authors: van den Pol AN , Kogelman L , Ghosh P , Liljelund P , Blackstone C
Published: April 2018

The expression of the metabotropic glutamate receptor mGluR1 was studied with Northern and Western blot analysis, with immunocytochemistry, and with Ca2+ digital imaging in the developing rat hypothalamus. mGluR1 is coupled to a G protein and activation by glutamate and related agonists leads to intracellular phosphotidylinositol hydrolysis and Ca2+ mobilization. mGluR1 RNA could be detected in embryonic hypothalamus, and by the day of birth and prior to the primary period of synaptogenesis, both mGluR1 RNA and protein were strongly expressed. In parallel experiments with digital imaging of cultured hypothalamic cells, some embryonic day 18 hypothalamic neurons and many astrocytes after 3 d in vitro showed Ca2+ responses to quisqualate and t-ACPD, and to glutamate in the absence of extracellular Ca2+. A greater number of embryonic neurons responded to NMDA than to agonists of the metabotropic receptor. With increased development time in culture, the number of neurons that responded to metabotropic glutamate receptor agonists increased. In the adult hypothalamus, mGluR1-immunoreactive neurons were widespread, and particularly dense in the dorsomedial, lateral, and anterior hypothalamus/preoptic areas, and in the mammillary body. Strongly immunoreactive cells were interspersed among neurons with no immunoreactivity. In developing neurons a diffuse immunostaining appeared along dendrites and somata. With time, beginning in the first week after birth, strongly stained puncta appeared, possibly associated with synaptic specializations. These puncta were numerous on dendrites of some adult neurons, and were the most strongly stained regions of neurons. Neurons developing in vitro at low neuron densities showed a development of mGluR1 immunoreactivity similar to that of neurons in vivo, but with a delayed progression of immunostaining. We found no obvious staining of axons or of astrocytes. A strong expression of mGluR1 protein was found in the hypothalamus during the first 2 postnatal weeks; this expression was partially reduced in adults. In contrast, cerebellum showed no reduction in mGluR1 protein in adults. Together these data suggest a complex regulation of mGluR1 during development, with sufficient expression of functional receptors in the developing hypothalamus to modulate morphogenesis and synaptogenesis, and later to play a role in transduction of glutamate signals in the adult. Different regions of the brain showed dramatic differences in the way each expresses mGluR1 during development.
mu-Opioid receptor-mediated reduction of neuronal calcium current occurs via a G(o)-type GTP-binding protein
Authors: Moises HC , Rusin KI , Macdonald RL
Published: April 2018

It has recently been shown that the activation of mu-opioid receptors inhibits several components of calcium channel current in rat DRG sensory neurons. mu-Opioid receptors, acting through the pertussis toxin (PTX)-sensitive substrate Gi, also reduce the activity of neuronal adenylate cyclase, but the relationship of this effect to changes in calcium channel activity has yet to be determined. Using whole-cell recordings from acutely isolated rat DRG neurons, we examined the ability of the mu-opioid-selective agonist Tyr-Pro-NMe-Phe- D-Pro-NH2 (PLO17) to reduce calcium current after treatment with PTX and in the presence of the nonhydrolyzable GTP analog guanosine 5′-[- thio]triphosphate (GTP gamma S), to assess the role of G-proteins in the coupling of mu-opioid receptors to calcium channels. Inhibition of current by PLO17 was mimicked or rendered irreversible by intracellular administration of GTP gamma S, an activator of G-proteins, and was blocked by pretreatment of neurons with PTX. In contrast, when the catalytic subunit of cAMP-dependent protein kinase was included in the recording pipette, calcium currents increased in magnitude throughout the recording without attenuation of responses to PLO17. Thus, the mu- opioid-induced inhibition of calcium current occurs through activation of a Gi- or G(o)-type G-protein, but independent of changes in adenylate cyclase activity. As a first step in identifying this G- protein, we compared the ability of several antisera directed against specific regions of Gi and G(o)alpha subunits to block the inhibition in current by PLO17. Intracellular dialysis with an antiserum specific for G(o) (GC/2) attenuated calcium current inhibition by PLO17 in five of six neurons by an average of 75%. In contrast, there was no attenuation in the response to PLO17 when neurons were dialyzed with an anti-Gi1 alpha/Gi2 alpha antiserum (AS/7) or antibodies specific for alpha subunits of Gi proteins (Gi1/Gi2 or Gi3) in an identical manner. These results suggest that in rat DRG neurons mu-opioid receptors couple to calcium channels via the PTX-sensitive G(o) subclass of GTP- binding proteins.
Laminar selectivity of the cholinergic suppression of synaptic transmission in rat hippocampal region CA1: computational modeling and brain slice physiology
Authors: Hasselmo ME , Schnell E
Published: April 2018

ACh may set the dynamics of cortical function to those appropriate for learning new information. In models of the putative associative memory function of piriform cortex, selective suppression of intrinsic but not afferent fiber synaptic transmission by ACh prevents recall of previous input from interfering with the learning of new input (Hasselmo, 1993). Selective cholinergic suppression may play a similar role in the hippocampal formation, where Schaffer collateral synapses in stratum radiatum (s. rad) may store associations between activity in region CA3 and the entorhinal cortex input to region CA1 terminating in stratum lacunosum-moleculare (s. l-m). A computational model of region CA1 predicts that for effective associative memory function of the Schaffer collaterals, cholinergic suppression of synaptic transmission should be stronger in s. rad than in s. l-m. In the hippocampal slice preparation, we tested the effect of the cholinergic agonist carbachol (0.01–500 microM) on synaptic transmission in s. rad and s. l-m. Stimulating and recording electrodes were simultaneously placed in both layers, allowing analysis of the effect of carbachol on synaptic potentials in both layers during the same perfusion in each slice. Carbachol produced a significantly stronger suppression of stimulus- evoked EPSPs in s. rad than in s. l-m at all concentrations greater than 1 microM. At 100 microM, EPSP initial slopes were suppressed by 89.1 +/- 3.0% in s. rad, but only by 40.1 +/- 4.1% in s. l-m. The muscarinic antagonist atropine (1 microM) blocked cholinergic suppression in both layers. These data support the hypothesis that synaptic modification of the Schaffer collaterals may store associations between activity in region CA3 and the afferent input to region CA1 from the entorhinal cortex. In simulations, feedback regulation of cholinergic modulation based on activity in region CA1 sets the appropriate dynamics of learning for novel associations, and recall for familiar associations.
An FCCP-sensitive Ca2+ store in bullfrog sympathetic neurons and its participation in stimulus-evoked changes in [Ca2+]i
Authors: Friel DD , Tsien RW
Published: April 2018

This study describes a Ca2+ store in fura-2-loaded bullfrog sympathetic neurons that modulates [Ca2+]i responses elicited by either depolarization or Ca2+ release from a caffeine- and ryanodine-sensitive store. This store is insensitive to caffeine and ryanodine, but is sensitive to the protonophore carbonyl cyanide p- trifluoromethoxyphenylhydrazone (FCCP). The FCCP-sensitive store slows both the rise in [Ca2+]i during stimulation (apparently by accumulating Ca2+ from the cytosol) and the recovery following stimulation (by releasing the accumulated Ca2+ into the cytosol). For a fixed level of depolarization, recovery is slowed to an extent that depends on stimulus duration. [Ca2+]i imaging shows that these effects are prominent in the soma but not in growth cones. Ca2+ uptake by the FCCP- sensitive store appears to be strongly [Ca2+]i dependent, since it becomes influential only when [Ca2+]i approaches approximately 500 nM. Therefore, this store may specifically influence [Ca2+]i during moderate and strong stimulation. The effect of the store on responses to depolarization can be accounted for by a simple three-compartment scheme consisting of the extracellular medium, the cytosol, and a single internal store with a [Ca2+]i-dependent uptake mechanism resembling the mitochondrial Ca2+ uniporter. The store's effect on responses to caffeine-induced Ca2+ release can be accounted for by including a second internal compartment to represent the caffeine- sensitive store. While the identity of the FCCP-sensitive store is unknown, its sensitivity to FCCP is consistent with a mitochondrial pool. It is suggested that by modulating the temporal properties of [Ca2+]i following stimulation, the FCCP-sensitive store may influence the degree of activation of intracellular [Ca2+]i-dependent processes.
Schwann cells induce sprouting in motor and sensory axons in the adult rat spinal cord
Authors: Li Y , Raisman G
Published: April 2018

Circumscribed lesions were made within either the corticospinal tract or the ascending dorsal column tracts at the upper cervical level in adult rats. The responses of the tract axons were studied by orthograde transport from injections of horseradish peroxidase or biocytin. At 2 d, the ends of the cut axons were swollen, and the lesions induced en passant varicosities in the adjacent uncut axons. Although there have been reports of retraction, we found that even after several weeks, large numbers of cut axons still persisted in the central lesion area (where there was complete tissue destruction and intense macrophage infiltration), and also in the adjacent regions of the tract. The cut ends were expanded into a variety of shapes--large, complex, bulbous, and recurved--and many had profuse local branches with or without small, terminal-type varicosities. A suspension of Schwann cells cultured from neonatal sciatic nerve was injected by a minimally traumatic air pressure microinjection technique so as to form a bolus, comparable in size to the lesions, in either the corticospinal or the ascending dorsal column tracts at the upper cervical level. Despite previous findings that corticospinal axons do not elongate into peripheral nerve grafts, we found that both corticospinal and ascending dorsal column axons sprouted in response to contact with the transplanted Schwann cells. The response to the Schwann cells was much more rapid than to the lesions. By 2 d, in both descending and ascending tracts, both the axons that had been severed at the time of injection and also the adjacent uncut axons had already given rise to the branches that (unlike the localized sprouting seen after the long- term lesions) extended for considerable distances parallel to and fasciculating with each other and with the uncut tract axons. In addition, a mass of fine, tortuous, varicose branches invaded the superficial parts of the Schwann cell grafts, where they formed aborizations with small bead-like expansions resembling presynaptic boutons; as in their normal terminal fields, the aborizations formed by the corticospinal axons were smaller and finer than those formed by the ascending axons.
Morphological evidence that hypothalamic substance P-containing afferents are capable of filtering the signal flow in the monkey hippocampal formation
Authors: Leranth C , Nitsch R
Published: April 2018

This study in the African green monkey (Cercopithecus aethiops) was designed to characterize the neurochemical features of hippocampal nonpyramidal neurons that are specific synaptic targets of substance P- containing projective neurons located in the supramammillary nucleus. Our previous studies provided evidence for an excitatory nature to this hypothalamo-hippocampal pathway and described the mode of termination of these afferents on hippocampal principal neurons. The present correlated light and electron microscopic immunocytochemical analysis, using the nickel-diaminobenzidine/diaminobenzidine double-labeling technique, revealed that this hippocampal afferent system establishes multiple, exclusively asymmetric synapses with three specific subpopulations of nonpyramidal cells: (1) a small portion of parvalbumin-containing basket cells located periodically in or adjacent to the granule cell layer of the dentate gyrus, which therefore inhibit only a subpopulation of granule cells; (2) some of the calbindin- immunoreactive local circuit neurons located in the hilar area; and (3) calbindin-positive cells occurring exclusively in the stratum molecular of the middle portion of the CA3 subfield. Postembedding studies revealed that the aforementioned calbindin-containing cells are GABAergic inhibitory neurons. Our studies indicate that hypothalamic afferents can effectively filter the information flow at different levels of the excitatory signal loop in the monkey hippocampal formation. Dentate granule cells, which are only stimulated by hypothalamic afferents, will transfer excitatory signals differently than those that are controlled by a feedforward inhibitory mechanism initiated by these fibers. In the CA3 subfield, the signal flow can again be depressed by those pyramidal neurons that are inhibited by calbindin-containing cells receiving an excitatory hypothalamic input.
Distributed processing of pain and vibration by the human brain
Authors: Coghill RC , Talbot JD , Evans AC , Meyer E , Gjedde A , Bushnell MC , Duncan GH
Published: April 2018

Pain is a diverse sensory and emotional experience that likely involves activation of numerous regions of the brain. Yet, many of these areas are also implicated in the processing of nonpainful somatosensory information. In order to better characterize the processing of pain within the human brain, activation produced by noxious stimuli was compared with that produced by robust innocuous stimuli. Painful heat (47–48 degrees C), nonpainful vibratory (110 Hz), and neutral control (34 degrees C) stimuli were applied to the left forearm of right-handed male subjects. Activation of regions within the diencephalon and telencephalon was evaluated by measuring regional cerebral blood flow using positron emission tomography (15O-water-bolus method). Painful stimulation produced contralateral activation in primary and secondary somatosensory cortices (SI and SII), anterior cingulate cortex, anterior insula, the supplemental motor area of the frontal cortex, and thalamus. Vibrotactile stimulation produced activation in contralateral SI, and bilaterally in SII and posterior insular cortices. A direct comparison of pain and vibrotactile stimulation revealed that both stimuli produced activation in similar regions of SI and SII, regions long thought to be involved in basic somatosensory processing. In contrast, painful stimuli were significantly more effective in activating the anterior insula, a region heavily linked with both somatosensory and limbic systems. Such connections may provide one route through which nociceptive input may be integrated with memory in order to allow a full appreciation of the meaning and dangers of painful stimuli. These data reveal that pain-related activation, although predominantly contralateral in distribution, is more widely dispersed across both cortical and thalamic regions than that produced during innocuous vibrotactile stimulation. This distributed cerebral activation reflects the complex nature of pain, involving discriminative, affective, autonomic, and motoric components. Furthermore, the high degree of interconnectivity among activated regions may account for the difficulty of eliminating pathological pain with discrete CNS lesions.
Activity-dependent expression and distribution of M1 muscarinic ACh receptors in visual cortex neuronal cultures
Authors: Wang Y , Gu Q , Mao F , Haugland RP , Cynader MS
Published: April 2018

The fluorescently labeled muscarinic M1 receptor-selective antagonist BODIPY FL pirenzepine has been employed to study the activity-dependent distribution and expression of muscarinic M1 ACh receptors (M1AChRs) in cultured neurons derived from rat visual cortex. Displacement experiments showed that like pirenzepine, binding of BODIPY FL pirenzepine was specific to M1 receptors and its K(i) was similar to that of unlabeled pirenzepine. Using confocal laser scanning microscopy, M1 receptors were predominantly localized to cell bodies early in development in the culture environment. After 2 weeks in culture, the receptors showed labeling not only in cell bodies but also in neuritic processes, especially on the initial segments of the processes. Chronic membrane depolarization with 40 mM potassium chloride caused a dramatic increase in M1 receptor expression on these neurons. Conversely, blockade of neuronal activity with 0.1 microM TTX decreased expression of the receptors. Receptor expression increased after cells were treated chronically with 50 nM pirenzepine, whereas it decreased after exposure to 10 microM carbachol. The results demonstrate for the first time the exact location of muscarinic receptors in living cultured neurons and also the activity-dependent expression of M1 receptors on these neurons. Both chronic membrane depolarization and antagonist application upregulate receptor expression, whereas blocking bioelectrical activity or chronic agonist application downregulates expression.
Role of phosphorylation in desensitization of acetylcholine receptors expressed in Xenopus oocytes
Authors: Hoffman PW , Ravindran A , Huganir RL
Published: April 2018

The nicotinic acetylcholine receptor (AChR) is a pentameric complex made up of four types of subunits in the stoichiometry alpha 2 beta gamma delta. These subunits have been shown to be differentially phosphorylated by cAMP-dependent protein kinase (PKA) protein kinase C, and a protein tyrosine kinase. A variety of studies have suggested that phosphorylation of the AChR in vitro and in vivo regulates the rate of desensitization of the receptor. In this study we have used site- specific mutagenesis and patch-clamp techniques to examine the role of phosphorylation in the regulation of desensitization of the AChR expressed in Xenopus oocytes Expression of wild-type AChR in Xenopus oocytes results in the constitutive phosphorylation of the AChR on the gamma and delta subunits. This phosphorylation is apparently due to the high basal level of PKA in oocytes since a specific peptide inhibitor of PKA completely eliminated phosphorylation of the AChR by oocyte extracts in vitro. The phosphorylation of the AChR in oocytes was not significantly enhanced by forskolin or cAMP analogs or by coexpression with the catalytic subunit of PKA, suggesting that the basal activity of PKA in oocytes is sufficient to phosphorylate the receptor to a high stoichiometry. Using site-specific mutagenesis, the sites of phosphorylation were determined to be serines 353 and 354 on the gamma subunit and serines 361 and 362 on the delta subunit. To examine the functional properties of wild-type and mutant receptors lacking phosphorylation sites, we used patch-clamp techniques to measure the responses of out-side-out patches to repetitive pulses of ACh using a rapid perfusion system. Wild-type and mutant receptors showed rapid concentration-dependent activation and desensitization to applied agonist. The time constant of desensitization of ensemble mean currents ranged from several hundred milliseconds at low ACh concentrations to 100–200 msec at saturating concentrations. The desensitization time constants for mutant receptors lacking all phosphorylation sites were significantly slower than wild-type phosphorylated receptors at all concentrations of ACh tested. In addition, mutant receptors that had the serine residues changed to glutamate residues in order to mimic the negative charge of the phosphorylated serine residue produced receptors that had desensitization rates approaching those of the wild-type phosphorylated receptor. These results provide further support that phosphorylation of the nicotinic ACh receptor regulates rate of desensitization.
Modulation of skeletal muscle sodium channels in a satellite cell line by protein kinase C
Authors: Numann R , Hauschka SD , Catterall WA , Scheuer T
Published: April 2018

Adult vertebrate skeletal muscle sodium channels are responsible for the spread of excitation from the end-plate through the muscle membrane and transverse tubular system that ultimately leads to contraction. These channels can be distinguished from other sodium channels by their sensitivity to both mu-conotoxin and TTX. The mouse satellite muscle cell line MM14 expresses only TTX- and mu-conotoxin-sensitive sodium channels having the physiological characteristics of adult skeletal muscle channels in both undifferentiated myoblasts and differentiated myotubes. Using undifferentiated and differentiated MM14 cells as well as primary cultures of rat skeletal muscle, we have examined modulation of adult skeletal muscle sodium channels by activators of protein kinase C (PKC). Stimulation of PKC by 1-oleoyl-2-acetyl-sn-glycerol (OAG) slows sodium current macroscopic inactivation rate by up to 70% and reduces the peak sodium current as much as 88%. Single-channel analysis reveals prolonged single channel openings and greatly increased probability of multiple channel openings during sustained depolarizations. These effects are due to PKC activation since they are blocked by a specific peptide inhibitor of PKC. The two effects of OAG are sequential. Low OAG concentrations can cause slowed macroscopic sodium current inactivation in the absence of peak current reduction, and intermediate concentrations of OAG cause slowing of inactivation followed by reduction of peak current. The separation of these two effects indicates that PKC modulation of the skeletal muscle sodium channel may occur by phosphorylation at two independent sites. PKC modulation of muscle sodium channels is expected to have important effects on muscle excitability and resultant contractile activity. Detection of adult skeletal muscle ion channels in replicating MM14 cells suggest that satellite cells may express a distinct subset of muscle-specific genes prior to activation of the terminal differentiation program.
Differential regulation of the release of the same peptide transmitters from individual identified motor neurons in culture
Authors: Whim MD , Lloyd PE
Published: April 2018

Aplysia motor neurons B1, B2, and B15 synthesize the small cardioactive peptides A and B (SCPs). In previous studies using semi-intact preparations we have demonstrated that the SCPs are released from B15. Significant peptide release only occurred when B15 was stimulated at a high frequency or at lower frequencies with a long burst duration. In the behaving animal B15 fires in patterns expected to release the SCPs. In contrast, in the behaving animal, neurons B1 and B2 fire at much lower frequencies. We therefore examined whether similar aspects of the stimulation pattern governed the release of the SCPs from B1 and B2. To monitor peptide release, all three neurons were individually cultured and newly synthesized peptides labeled with 35S-methionine. Release of labeled SCPs was detected by HPLC of extracts of superfusates. By keeping spike number constant and varying the stimulation pattern, the release of the SCPs from B1 and B2 was found to be pattern insensitive. That is, regardless of the stimulation paradigm, each action potential released a similar amount of peptide. By severing the primary neurite, peptide release was found to occur mainly from the regenerated neurites and most likely from the intensely immunoreactive varicosites. Calcium- and stimulation-dependent release of the SCPs and a third neuropeptide termed buccalin A from motor neuron B15 was also observed in culture. The release of these peptides from B15 was found to be pattern sensitive as was observed in the semi-intact preparations.(ABSTRACT TRUNCATED AT 250 WORDS)
Gradual tolerance of metabolic activity is produced in mesolimbic regions by chronic cocaine treatment, while subsequent cocaine challenge activates extrapyramidal regions of rat brain
Authors: Hammer RP , Cooke ES
Published: April 2018

Acute administration of cocaine is known to enhance extracellular dopamine levels in the striatum and to activate immediate-early gene expression in striatal neurons. Regional cerebral metabolic rate for glucose (rCMRglc) reportedly increases in extrapyramidal and mesolimbic brain regions in response to acute cocaine treatment. However, chronic administration attenuates the cocaine-induced enhancement of regional dopamine response and the induction of immediate-early gene expression in these regions. Chronic treatment also produces tolerance to cocaine's reinforcing effects. Thus, differential responses to cocaine occur with increasing length of treatment. Therefore, we examined the time course of effects of repeated daily cocaine treatment on rCMRglc in rat brain. Acute administration of 10 mg/kg cocaine slightly increased rCMRglc in mesolimbic and extrapyramidal regions. However, no significant effects were observed until more than 7 d of treatment, whereupon rCMRglc was reduced compared to saline treatment in the infralimbic portion of the medial prefrontal cortex, nucleus accumbens, olfactory tubercle, habenula, amygdala, and a few other brain regions. In contrast, after 13 d of 10 mg/kg cocaine treatment, challenge with 30 mg/kg cocaine increased rCMRglc in the striatum, globus pallidus, entopeduncular nucleus, subthalamus, substantia nigra pars reticulata, and a few other regions without affecting limbic or mesolimbic regions. Thus, repeated daily treatment with a low dose of cocaine gradually decreased metabolic activity particularly in mesolimbic regions. Subsequent treatment with a higher dose produced metabolic activation mostly in extrapyramidal regions. This effect of chronic treatment could represent tolerance to the initial metabolic response, which can be replicated thereafter but only by increasing the drug dose. These results suggest that tolerance to the metabolic effects of cocaine in selective mesolimbic circuits may contribute to the development of behavioral dependence with repeated exposure.
Chronic electroconvulsive seizure (ECS) treatment results in expression of a long-lasting AP-1 complex in brain with altered composition and characteristics
Authors: Hope BT , Kelz MB , Duman RS , Nestler EJ
Published: April 2018

Gene transcription is likely to play a role in the biochemical adaptations thought to underlie the long-term behavioral changes observed following various chronic treatments. The AP-1 (activator protein-1) complex is a well-studied transcription factor capable of regulating gene transcription. We therefore examined the regulation of the AP-1 complex in rat cerebral cortex and hippocampus following electroconvulsive seizures (ECS), known to induce biochemical alterations in the brain after chronic treatment. We show that 10 d of chronic ECS treatment results in an AP-1 binding complex that persists for at least 7 d in the cortex and hippocampus. In contrast, AP-1 binding returns to control levels within 18 hr of a single acute ECS. Supershift experiments and Western blots show that the chronic AP-1 complex contains two novel Fos-related antigens (Fras) of 35 and 37 kDa that do not appear following a single acute ECS. The chronically induced 35 and 37 kDa Fras and the chronic AP-1 complex show similar time courses for induction by repeated ECS. Moreover, the 37 kDa Fra band persists for at least 7 d following chronic ECS treatment, as observed for the chronic AP-1 complex. Competition experiments indicate that the relative affinities of the acute and chronic AP-1 complexes for several AP-1-like sites are similar, although there was approximately a twofold difference in the affinity for one particular AP-1-like site. The altered composition of the chronic AP-1 complex, and differences in half-life, DNA binding affinity, and possibly transcriptional activating properties are likely to cause changes in the overall pattern of gene expression, which may underlie some of the long-term biochemical adaptations observed following chronic ECS and other chronic perturbations.
Opioid receptor activation is one factor underlying the frequency dependence of mossy fiber LTP induction
Authors: Derrick BE , Martinez JL
Published: April 2018

The contribution of high-frequency synaptic activity to the induction of long-term potentiation (LTP) in the opioid peptide-containing mossy fiber projection was investigated in vivo in anesthetized rats. Because high-frequency mossy fiber activity is essential for both the release of opioid peptides and the induction of mossy fiber LTP, we investigated whether the activation of opioid receptors underlies the requirement of sustained high-frequency mossy fiber activity for LTP induction. Mossy fiber responses were found to have a distinct threshold for the number of 100 Hz pulses necessary to induce LTP, with bursts of 25–30 pulses being the minimum for LTP induction. Application of 1 nmol of the mu-opioid receptor agonist DAMGO to the CA3 region potentiated mossy fiber responses, but, unlike for mossy fiber LTP, this potentiation could be reversed by mu-opioid receptor antagonist CTOP. Stimulation of the mossy fibers with either a single burst of 15 pulses at 100 Hz or application of 100 pmol of DAMGO was ineffective in potentiating mossy fiber responses. However, delivery of a 15 pulse burst 10 min following DAMGO application was effective in potentiating mossy fiber responses. This potentiation was not reversed by CTOP and it occluded stimulation-induced LTP, suggesting that brief bursts delivered in the presence of DAMGO had induced mossy fiber LTP. The release of opioid peptides and the resulting activation of mu-opioid receptors is suggested as one factor that underlies the requirement of sustained high-frequency stimulation for the induction of mossy fiber LTP.(ABSTRACT TRUNCATED AT 250 WORDS)
Enhancement of Ca current in the accessory radula closer muscle of Aplysia californica by neuromodulators that potentiate its contractions
Authors: Brezina V , Evans CG , Weiss KR
Published: April 2018

A major goal of neuroscience is to identify the neural and cellular mechanisms of behavior and its plasticity. Progress toward this goal has come particularly from work with a small number of tractable model preparations. One of these is the simple neuromuscular circuit consisting of the accessory radula closer (ARC) muscle of the mollusk Aplysia californica and its innervating motor and modulatory neurons. Contraction of the ARC muscle underlies a component of Aplysia feeding behavior, and plasticity of the behavior is in large part due to modulation of the amplitude and duration of the contractions of the muscle by a variety of modulatory neurotransmitters and peptide cotransmitters, among them the small cardioactive peptides (SCPs), myomodulins (MMs), and serotonin (5-HT). We have studied single dissociated ARC muscle fibers in order to determine whether modulation of membrane ion currents in the muscle might underlie these effects. First, we confirmed that the dissociated fibers were functionally intact: just as with the whole ARC muscle, their contractions were potentiated by 5-HT and SCPB and potentiated as well as depressed by MMA, and their cAMP content was greatly elevated by 5-HT, SCPA and SCPB, and to a lesser extent by MMA and MMB. Next, using voltage-clamp techniques, we found that two ion currents present in the fibers were indeed modulated. The fibers possess a dihydropyridine-sensitive, high- threshold “L”-type Ca current. This current was enhanced by the modulators that potentiate ARC-muscle contractions--5-HT, SCPA and SCPB, and MMA and MMB--but not by buccalinA, a modulator that does not act directly on the ARC muscle. All of the potentiating modulators, as well as elevation of cAMP in the fibers by forskolin or a cAMP analog, maximally enhanced the current about twofold and mutually occluded each other's effects. Since the Ca current supplies Ca2+ necessary for contraction of the muscle, the enhancement of the current is a good candidate to be a major mechanism of the potentiation of the contractions. In the following article we report that the modulators also, to different degrees, activate a distinctive K current and thereby depress the contractions. Net potentiation or depression then depends on the balance between the relative strengths of the modulation of the two ion currents.