bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of long-term haloperidol treatment on the responsiveness of accumbens neurons to cholecystokinin and dopamine: electrophysiological and radioligand binding studies in the rat
Authors: Debonnel G , Gaudreau P , Quirion R , de Montigny C
Published: April 2018

Cholecystokinin (CCK) and dopamine (DA) coexist in a subpopulation of neurons of the ventral tegmental area projecting to the nucleus accumbens. The present experiments were undertaken to determine the effect of acute and long-term administration of haloperidol on the responsiveness of accumbens neurons to microiontophoretic applications of the sulfated cholecystokinin octapeptide (CCK-8S), kainate (KA), and DA and on the density of CCK, D1, and D2 receptors determined by radioautography. Acute administration of haloperidol (1 mg/kg, i.v.) did not modify the neuronal responsiveness to DA and KA but increased that to CCK-8S. Long-term treatment with haloperidol decanoate (4 mg/kg/week, i.m., for 3–5 weeks) induced a marked increase in the responsiveness to CCK-8S, without noticeable change of that to DA and KA. After a 5 week treatment, significant increases in the amounts of CCK and D2 binding were found in the nucleus accumbens, whereas D1 binding parameters remained unchanged. Since long-term haloperidol treatment results in a depolarization inactivation of A10 dopaminergic neurons, these results suggest that, despite the reduced firing activity of mesolimbic dopaminergic neurons induced by the long-term haloperidol treatment, dopamine is still released in an amount sufficient to maintain a normal neuronal responsiveness of postsynaptic accumbens neurons to DA, whereas the release of CCK is possibly decreased to a greater extent, resulting in an enhanced responsiveness of the neurons to this peptide.
Intracellular and surface acetylcholine receptors during the normal development of a frog skeletal muscle
Authors: Goldfarb J , Cantin C , Cohen MW
Published: April 2018

125I-alpha-bungarotoxin (125I-alpha BT) was used to measure the pool sizes of surface and intracellular acetylcholine receptors (AChRs) in the myotomal muscle of Xenopus laevis over a developmental period (stages 23–48; 1.03–7.5 d) which ranged from initial to mature stages of neuromuscular synaptogenesis. The surface pool increased progressively throughout development. The intracellular pool increased more slowly and also underwent a transient decrease. Linear regression indicated that AChRs begin to appear intracellularly and in the surface membrane at embryonic ages of 13.2 and 18.5 hr, respectively. The findings also suggest that newly synthesized AChRs contribute much more to the intracellular pool than do AChRs internalized from the surface membrane and that the rates of supply and/or intracellular resident times of these 2 sources of intracellular AChRs change during the course of normal development. Carbamylcholine, even at concentrations 10-fold greater than needed to block completely 125I-alpha BT binding to surface AChRs, blocked specific intracellular binding by only 80%. Considered in the light of previous studies on cell cultures, these results suggest that 20% of the intracellular sites are on alpha- subunits not yet assembled into pentameric AChRs. Light microscope radioautography revealed an essentially uniform distribution of intracellular AChRs along the length of the muscle cells. It is concluded that during the normal development of Xenopus myotomal muscle the accumulation and maintenance of AChRs in the postsynaptic membrane occurs in the absence of any preferential concentration of intracellular AChRs in the subsynaptic region.
Firing patterns and synaptic potentials of identified giant aspiny interneurons in the rat neostriatum
Authors: Wilson CJ , Chang HT , Kitai ST
Published: April 2018

Intracellular recordings were made in vivo from 9 giant aspiny neurons in the neostriatum of urethane-anesthetized rats. The cells were identified by intracellular staining with HRP or biocytin. The neurons exhibited morphological features typical of neostriatal cholinergic interneurons. Six of the cells were obtained from intact animals, while 3 were recorded from rats with ipsilateral hemidecortications. Giant aspiny neurons were characterized by their slow irregular but tonic (3–10/sec) spontaneous activity and long-duration action potentials. Examination of the underlying membrane potential trajectories during spontaneous firing revealed that individual action potentials were triggered from spontaneous small (1–5 mV) depolarizing potentials. These spontaneous potentials exhibited the voltage sensitivity of ordinary EPSPs. They were much less frequent during the 80–200 msec pause in tonic afferent input that follows the excitation evoked by cortical or thalamic stimulation, and were decreased in frequency in decorticate animals. Their rise times and half-widths matched those expected for unitary synaptic potentials placed proximally on the surface of the neurons. Low-intensity stimulation of neostriatal afferents produced small short-latency EPSPs that appeared to be composed of responses identical to the spontaneous depolarizing potentials. The latencies of the EPSPs evoked from the cerebral cortex and thalamus were consistent with a monosynaptic input from both structures, but the maximal size of the EPSPs was much smaller than that evoked in spiny neurons, suggesting that a smaller number of afferent inputs make synapses with each of the aspiny cells. Giant aspiny neurons exhibited much larger input resistances and longer time constants than spiny neostriatal neurons. They also exhibited relatively linear steady-state current-voltage relationship compared to spiny projection cells. Input resistances ranged from 71–105 M omega, and time constants ranged from 17.8–28.5 msec. Analysis of the charging transients in response to current pulses yielded estimates of dendritic length of approximately 1 length constant. Repetitive firing of the neurons was limited by a powerful spike afterhyperpolarization and by a strong spike frequency adaptation. The sensitivity of the giant aspiny interneuron to a relatively small number of proximal afferent synaptic contacts, its tonic firing, and its widespread dendritic and axonal fields place it in an excellent position to act as a modulator of the excitability of neostriatal projection neurons in advance of the onset of movement-related neostriatal activity.
Localization and characterization of 35S-t- butylbicyclophosphorothionate binding in rat brain: an autoradiographic study
Authors: Edgar PP , Schwartz RD
Published: April 2018

35S-t-butylbicyclophosphorothionate (TBPS) binding to slide-mounted rat brain sections was characterized for subsequent autoradiographic analysis. Cortical brain mash slices, preincubated with EDTA to remove endogenous GABA, were used for biochemical characterization. Steady state for 35S-TBPS binding was reached by 3 hr of incubation at 22 degrees C. The association rate constant (K1) and dissociation rate constant (K2) were 0.377 min-1 microM-1 and 0.011 min-1, respectively. Dissociation was monophasic and slow (t1/2 = 80 min). The kinetically derived KD was 29.4 nM. Scatchard analysis indicated a single population of binding sites with a KD of 21.0 +/- 2.2 nM and a Bmax of 1.59 +/- 0.13 pmol/mg protein. Both picrotoxin and muscimol inhibited 35S-TBPS binding completely with IC50s of 251 +/- 13 nM and 203 +/- 41 nM and nHs of 0.98 and 1.4, respectively. The distribution of 35S-TBPS binding sites in the rat brain resembles that of other ligands that bind to GABAA receptor complex with some regionally specific differences. Regions with a high degree of 35S-TBPS binding included the inferior colliculus, medial septal nucleus, central and paracentral nuclei of the thalamus, olfactory tubercle, zona incerta, dentate gyrus, and substantia nigra. 35S-TBPS preferentially bound to the molecular vs granular layer of the cerebellum. Omission of the preincubation markedly but variably decreased 35S-TBPS binding. The greatest regional decreases occurred in areas with a high degree of GABA synthesis. In addition, 35S-TBPS binding was inhibited to different degrees in the cell layers of the cerebellum. The addition of 1 microM GABA to the incubation medium of preincubated slices also produced variable decreases in 35S-TBPS binding to cerebellar layers. These findings support previous studies that demonstrate GABAA receptor heterogeneity. Our study confirms the suitability of 35S-TBPS for use as a ligand in autoradiography and demonstrates that the distribution of 35S-TBPS binding sites is significantly influenced by the preincubation-incubation conditions used.
Optokinetic stimulation increases corticotropin-releasing factor mRNA in inferior olivary neurons of rabbits
Authors: Barmack NH , Young WS
Published: April 2018

Stimulus-specific changes in levels of corticotropin-releasing factor (CRF) mRNA in inferior olivary neurons were studied in unanesthetized rabbits. The possible functional importance of CRF, a neuropeptide that is expressed in all inferior olivary neurons, in cerebellar synaptic transmission was investigated in a subpopulation of inferior olivary neurons, the caudal dorsal cap. Previous studies have shown that unidirectional, binocular optokinetic stimulation increases the level of discharge in neurons located in one of the caudal dorsal caps of the inferior olive and decreases the level of discharge in neurons in the opposite dorsal cap. We investigated the influence of prolonged (1–144 hr), unidirectional, binocular, optokinetic stimulus on the levels of CRF mRNA in dorsal cap neurons, measured with the technique of hybridization histochemistry. Rabbits were placed at the center of a cylindrical optokinetic drum that rotated at a constant velocity of 5 deg/sec, stimulating one eye in the posterior-to-anterior direction and the contralateral eye in the anterior-to-posterior direction. Posterior- to-anterior stimulation of the left eye evoked increased activity of inferior olivary neurons located in the right caudal dorsal cap. Conversely, anterior-to-posterior stimulation of the right eye evoked decreased activity of neurons in the left caudal dorsal cap. The levels of CRF mRNA in dorsal cap neurons that were activated by prolonged optokinetic stimulation were increased by a factor of 4 to 7 after 48 hr of stimulation and by more than a factor of 10 after 144 hr of optokinetic stimulation. These optokinetically induced increases in CRF mRNA decayed to background levels after 30 hr. Our observations implicate CRF in visual olivocerebellar function and suggest that CRF may play a role in the plastic control of eye movement mediated by the visual olivocerebellar system.
Multiple nicotinic acetylcholine receptor genes are expressed in goldfish retina and tectum
Authors: Cauley K , Agranoff BW , Goldman D
Published: April 2018

cDNAs encoding a novel nAChR structural subunit (GFn alpha-3) and a ligand-binding subunit (GF alpha-3) have been isolated from a goldfish retina cDNA library. The protein encoded by GFn alpha-3 shares 88% amino acid similarity with that encoded by GFn alpha-2, a structural subunit gene previously identified to be expressed in this system (Cauley et al., 1989). The ligand-binding subunit (GF alpha-3) is likely the goldfish homolog of the rat alpha-3 gene (Boulter et al., 1986). Northern blots and S1 protection experiments show that GFn alpha- 3 and GF alpha-3 genes are expressed in retina and brain. GFn alpha-3 identifies multiple RNAs differing in their 3′ untranslated regions. In situ hybridization analysis demonstrates GFn alpha-3, GFn alpha-2, and GF alpha-3 expression by cells of the retinal ganglion cell layer. Unlike GFn alpha-2 and GF alpha-3, GFn alpha-3 is expressed at highest levels by cells of the retina's inner nuclear layer. In the optic tectum, both GF alpha-3 and GFn alpha-3 genes are expressed by cells of the periventricular zone, as well as more superficial layers. These results suggest the presence of multiple nAChR systems in retina and tectum. In addition, they indicate that tectal nAChRs may arise from remote (ganglion cell) as well as local (tectal cell) synthesis.
Non-NMDA receptor-mediated neurotoxicity in cortical culture
Authors: Koh JY , Goldberg MP , Hartley DM , Choi DW
Published: April 2018

The neurotoxicity of 3 non-NMDA glutamate receptor agonists--kainate, alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA), and quisqualate--was investigated quantitatively in dissociated murine cortical cultures. Five minute exposure to 500 microM kainate, but not AMPA, produced widespread acute neuronal swelling. Kainate-induced swelling was resistant to 2-amino-5-phosphonovalerate (APV) or replacement of extracellular sodium with choline but attenuated by either kynurenate or low concentrations of quisqualate. Unlike NMDA agonists, kainate or AMPA did not produce much late neuronal loss after a 5 min exposure. In contrast, 5 min exposure to 500 microM quisqualate produced both acute neuronal swelling and widespread late neuronal degeneration. This acute swelling was blocked by APV or by replacement of extracellular sodium by choline, consistent with mediation by NMDA receptors; we speculate that high concentrations of quisqualate may directly activate NMDA receptors or induce the release of endogenous glutamate. Quisqualate-induced late neuronal degeneration may be due to another unexpected process: cellular quisqualate uptake and delayed release, converting brief addition into prolonged exposure. Hours after thorough washout of exogenously added quisqualate, micromolar concentrations could be detected in the bathing medium by high performance liquid chromatography. With lengthy exposure (20–24 hr), all 3 non-NMDA agonists were potent neurotoxins, able to destroy neurons with EC50's of about 20 microM for kainate, 4 microM for AMPA, and 1 microM for quisqualate. Kynurenate and 6-cyano-7-nitroquinoxaline- 2,3-dione (CNQX), but not APV or L-glutamate diethyl ester, were effective in attenuating the neuronal degeneration induced by these agonists. CNQX was about 3 times more selective than kynurenate against kainate-induced neuronal injury, but CNQX was still nearly equipotent with APV against NMDA-induced injury. Gamma-D-glutamylaminomethyl sulfonate exhibited partial antagonist specificity for AMPA-induced toxicity.
Making arm movements within different parts of space: dynamic aspects in the primate motor cortex
Authors: Caminiti R , Johnson PB , Urbano A
Published: April 2018

The activity of 176 individual cells in the arm area of motor cortex (areas 4 and 6) was studied while monkeys made arm movements of similar direction within different parts of extrapersonal space. The behavioral paradigm used was a 3-dimensional reaction-time task aimed at dissociating the direction of movement, which remained similar across the work space, from the patterns of muscular activity and the angular joint excursions necessary to perform these movements. In agreement with other studies (Georgopoulos et al., 1982; Schwartz et al., 1988), we found that, within a given part of space, the activity of 169 (96.0%) cells studied increased most for a given preferred direction and less for other directions of movement. This change was graded in an orderly fashion. We further analyzed the orientation in space of the cells' preferred directions under the differing conditions of the task. We found that, as movements with similar trajectories were made within different parts of space, the cells' preferred directions changed spatial orientation. This change was of different magnitudes for different cells, but at the level of the population, it followed closely the changes in orientation of the arm necessary to perform the movements required by the task. Movement population vectors (Georgopoulos et al., 1983, 1986, 1988) computed from cell activity proved to be good predictors of movement direction regardless of where in space the movements were performed. These results indicate that motor cortical cells can code direction of movement in a way which is dependent on the position of the arm in space. The data are discussed in relation to the existence of mechanisms which facilitate the transformation between extrinsic and intrinsic coordinates. These transformations are necessary to perform arm movements to visual targets in space.
Basic fibroblast growth factor: receptor-mediated internalization, metabolism, and anterograde axonal transport in retinal ganglion cells
Authors: Ferguson IA , Schweitzer JB , Johnson EM
Published: April 2018

Basic fibroblast growth factor (bFGF) was radiolabeled and used in axonal transport studies to determine whether certain neuronal populations express functional receptors for bFGF. Unlike 125I-NGF, 125I-bFGF was not retrogradely transported in the adult rat sciatic nerve or from iris to trigeminal ganglion or superior cervical ganglion. However, after intraocular injection of 125I-bFGF into the posterior chamber of the eye of adult rats, radioactivity was detected within the retinal ganglion cell projections. This radioactivity was localized to the ipsilateral optic nerve and in the contralateral lateral geniculate body and the contralateral superior colliculus by using autoradiographic techniques. Direct measurement of the radioactivity in dissected brain regions was used to study the process of 125I-bFGF uptake and transport by retinal ganglion cells. The uptake and transport were specific for biologically active bFGF since neither denatured, biologically inactive 125I-bFGF nor 125I-NGF was taken up and transported. The uptake and transport of 125I-bFGF were saturable phenomena since they were blocked in the presence of excess, unlabeled bFGF. Wheat germ agglutinin, but not heparinase, blocked uptake and transport of 125I-bFGF, a finding that is consistent with the uptake being mediated by high-affinity bFGF receptors. Radioactivity from 125I- bFGF was transported in retinal ganglion cell axons in an anterograde direction at a maximum rate in excess of 1.7 mm/hr. No specific retrograde transport of bFGF to the retina was detected after 125I-bFGF was injected into the superior colliculus. The radioactivity from 125I- bFGF that accumulated in the superior colliculus was lost from this tissue with a half-life of about 22 hr. Autoradiography of proteins separated by SDS-PAGE demonstrated that 125I-bFGF was not substantially degraded in the retina after internalization within retinal ganglion cells. During anterograde transport, however, 125I-bFGF underwent limited proteolytic cleavage resulting in 3 prominent 125I-bFGF derivatives of molecular weights greater than 7000 Da. Although these were the major radioactive species recovered from the superior colliculus after intraocular injection, some intact 125I-bFGF was also detected within the innervated target. These results indicate that retinal ganglion cells express high-affinity receptors for bFGF, that these receptors mediate the internalization of bFGF, that internalized bFGF undergoes limited proteolytic cleavage, and that bFGF and its derivatives are anterogradely transported to the lateral geniculate body and the superior colliculus. These data raise the possibility that bFGF or its derivatives may act as an anterograde trophic factor in the visual system, a system that is known to undergo anterograde transneuronal cell death.
A comparison of peripheral and central axotomy effects on neurofilament and tubulin gene expression in rat dorsal root ganglion neurons
Authors: Wong J , Oblinger MM
Published: April 2018

The expression of major cytoskeletal protein mRNAs was studied in adult rat dorsal root ganglion (DRG) neurons after crushing either their central or peripheral branch axons. mRNA levels in DRG neurons were examined by quantitative in situ hybridization with radiolabeled cDNA probes specific for the low-molecular-weight neurofilament protein (NF- L) and beta-tubulin. The large-sized (greater than 1000 microns 2) neurons which give rise to myelinated axons in lumbar ganglia (L4 and L5) were studied 1 d through 8 weeks after either dorsal root or sciatic nerve crush. NF-L and beta-tubulin mRNA levels in axotomized DRG neurons were compared to those in contralateral control DRG neurons, as well as to those in normal (completely untreated) DRG cells. In the case of NF-L mRNA, changes were observed after central as well as peripheral branch axotomy and the time course and magnitude of changes were similar after both types of axotomy. NF-L mRNA levels initially decreased (first 2 weeks after crush) and then began to return towards control levels at longer survival times. Similar, but less pronounced, changes in NF-L mRNA levels also occurred in contralateral DRG neurons (which were uninjured); the changes in contralateral neurons were not simply a result of surgical stress since no changes in NF-L mRNA levels were observed in sham-operated DRG neurons. In the case of tubulin mRNA, changes were observed after central as well as peripheral branch axotomy by in situ hybridization, but the time course and magnitude of changes were different after each type of axotomy.(ABSTRACT TRUNCATED AT 250 WORDS)
Dihydropyridine actions on calcium currents of frog sympathetic neurons
Authors: Jones SW , Jacobs LS
Published: April 2018

Dihydropyridines (DHPs) generally have little effect on whole-cell calcium currents of neurons, even at concentrations far higher than those effective on muscle. Either neuronal calcium currents are much less sensitive to DHPs, or only a small proportion of the current is DHP-sensitive. We find that DHP agonists and antagonists act at low concentration on calcium currents in frog sympathetic neurons but that the effects are small even at optimal concentrations. The half-maximal dose (EC50) of the agonist Bay K 8644 is approximately 50 nM, and the effect of Bay K 8644 is blocked by 50% at approximately 300 nM nifedipine, from a holding potential of -80 mV. Nifedipine is more effective from a holding potential of -50 mV. These results suggest the presence of an L-type calcium current, with DHP sensitivity similar to L-currents in cardiac muscle. The predominant (greater than 90%) calcium current in frog sympathetic neurons is a DHP-resistant N-type current. However, high concentrations of DHPs (10 microM) partially block N-type calcium current, as well as voltage-dependent sodium and potassium currents.
Voltage-dependent nonlinearities in the membrane of locust nonspiking local interneurons, and their significance for synaptic integration
Authors: Laurent G
Published: April 2018

Some aspects of the electronic and active membrane properties of nonspiking local interneurons were studied in isolated locust thoracic ganglia, using the switched current- and voltage-clamp techniques in neuropilar recordings. The average transmembrane potential (Vr) of the interneurons was -58 +/- 6mV (n = 85), and the input resistance (in the linear region of the current-voltage curve) was 16.5 +/- 8 M omega (n = 19, range 8 to 32 M omega). The membrane and equalizing time constants were estimated from charging curves evoked by the injection of low density hyperpolarizing current pulses from about -80 mV, i.e., from voltages in the linear region of the I-V curve. The curves yielded 2 time constants (tau m and tau l) whose average values were 33.2 +/- 9 msec and 3.3 +/- 1 msec (n = 18), respectively. The mean specific membrane resistance is therefore about 33 k omega.cm2, assuming that the membrane capacitance is ca. 1 microF.cm-2. An outward rectification was always observed upon depolarization from potentials more negative than Vr and was accompanied by a decrease in input resistance and membrane time constant. The “resting” membrane, for example, had a time constant of 26.4 +/- 8 msec (n = 31). This outward rectification was due to at least 2 conductances with different inactivation kinetics, similar to the transient “A” and “delayed-rectifier” potassium conductances. No inward rectification was ever observed upon injection of hyperpolarizing current. In about 60% of the recordings, an active and TTX-resistant depolarizing process could be evoked by rapid depolarization around Vr. The voltage-dependent properties of the membrane of the nonspiking local interneurons had dramatic effects on the shape and time course of natural or evoked unitary PSPs. The half- width of EPSPs, for example, decreased by a factor of 7.5 if the membrane potential was shifted from -93 to -50 mV. When the membrane potential of an interneuron was altered with a triangular current waveform, the reduction of tonically occurring IPSPs depended more on the sign and rate of the induced voltage change than on the absolute transmembrane potential. For 2 identical instantaneous values of membrane potential, for example, the reduction of the PSPs was greater during the depolarizing phase than during the hyperpolarizing phase of the current waveform. The possible nature of the active membrane conductances underlying the nonlinear electrical behavior of the membrane is discussed, together with their functional significance for local circuit synaptic integration.
Regulation of short-term associative memory by calcium-dependent protein kinase
Authors: Matzel LD , Lederhendler II , Alkon DL
Published: April 2018

Neural and behavioral correlates of an associative memory in Hermissenda were examined during induction and/or formation of the memory. Hermissenda received either light (conditioned stimulus or CS) and rotation (unconditioned stimulus or US) paired (i.e., Pavlovian conditioning), light and rotation unpaired (pseudoconditioning), or no exposure to light and rotation. Following 9 pairings in a 6 min session, conditioned animals exhibited a contraction of the foot in response to a test CS presented 2 min after the last conditioning trial, whereas pseudoconditioned and untreated animals exhibited a foot extension to the same CS. In addition, both an associative and a nonassociative reduction in light-induced locomotion was observed. To examine neural correlates of this learning within minutes of acquisition, the isolated nervous system of the Hermissenda (containing the visual and vestibular organs) was trained with stimulus conditions identical to those used for the intact animal. Prior isolation and preparation of the nervous system permitted immediate intracellular recording following the final conditioning trial. Relative to pseudoconditioned and untreated animals, the B photoreceptors in conditioned nervous systems were found to have elevated input resistance (inversely related to K+ channel conductance and positively related to excitability) and exhibited increased steady-state depolarization in response to the light CS, as well as a prolonged depolarization after the CS offset. These neural correlates of the associative memory were attenuated if the protein kinase inhibitor H7 was present in the extracellular bath during conditioning, demonstrating in the reduced preparation that antagonism of protein kinase activity blocks the induction of membrane alterations of identified neurons that correlate with memory storage.(ABSTRACT TRUNCATED AT 250 WORDS)
Opiate withdrawal and the rat locus coeruleus: behavioral, electrophysiological, and biochemical correlates
Authors: Rasmussen K , Beitner-Johnson DB , Krystal JH , Aghajanian GK , Nestler EJ
Published: April 2018

We have compared the time course of the behavioral manifestations of opiate withdrawal to the in vivo activity of locus coeruleus (LC) neurons and to increases in the levels of G-proteins, adenylate cyclase, and cAMP-dependent protein kinase known to occur in the LC in opiate-dependent animals. Rats were given morphine by daily subcutaneous implantation of morphine pellets for 5 d. On the sixth day, morphine withdrawal was induced by subcutaneous administration of naltrexone, an opiate receptor antagonist, with additional doses given 6 and 24 hr later, conditions that resulted in sustained, maximal levels of withdrawal over the duration of the experiment. We found a striking parallel between the time courses of the behavioral signs and the increased activity of LC neurons during withdrawal, both of which appeared to follow 2 phases. There was an early, rapid phase, during which withdrawal signs and increased LC activity became most pronounced within 15–30 min after naltrexone administration, and then recovered rapidly by over 50% within 4 hr of withdrawal. Subsequently, there was a slower phase, during which the persisting withdrawal signs and elevated LC activity remained roughly constant from 4 to 24 hr and did not recover completely until after 72 hr of continuous withdrawal. Adenylate cyclase and cAMP-dependent protein kinase activities in isolated LC subcellular fractions, both elevated in dependent animals, recovered to control levels after 6 hr of withdrawal, in parallel with the rapid phase of withdrawal. Levels of G1 and Go, also elevated in dependent animals, remained only slightly elevated at 6 hr and returned to normal by 24 hr. Taken together, these data suggest that increased neuronal activity in the LC is associated temporally with the behavioral morphine withdrawal syndrome and that increased levels of G- proteins and an up-regulated cAMP system may contribute to the early withdrawal activation of these neurons.
Functional characteristics and sites of gene expression of the alpha 1, beta 1, gamma 2-isoform of the rat GABAA receptor
Authors: Malherbe P , Sigel E , Baur R , Persohn E , Richards JG , Mohler H
Published: April 2018

GABAA receptors, the major synaptic targets for the neurotransmitter GABA, constitute gated chloride channels. By their allosteric, drug- induced modulation, they serve as control elements for the regulation of anxiety, vigilance, and epileptiform activity. The structural requirements of fully functional GABAA receptors in the mammalian brain have remained elusive so far. We report here on the cloning of the gamma 2-subunit cDNA of rat brain and its functional analysis by coexpression with the alpha 1- and beta 1-subunits in Xenopus oocytes, and on the sites of gene expression of the 3 subunits in the rat brain. The recombinant receptor displayed GABA-inducible currents (Imax = 6 microA; Ka = 75 microM) which were allosterically modulated by benzodiazepine receptor ligands (enhancement and inhibition by diazepam and methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate, respectively). In the absence of GABA, pentobarbital elicited a maximal current amplitude similar to that of GABA. A minor population of channels is expressed which is open in the absence of GABA or pentobarbital. Mapping subunit gene expression by in situ hybridization histochemistry suggests that the alpha 1-, beta 1-, and gamma 2- subunits are likely receptor constituents in some neuronal populations, e.g., mitral cells of the olfactory bulb, pyramidal cells of the hippocampus, and granule cells of the dentate gyrus and cerebellum.
Site-specific phosphorylation of the middle molecular weight human neurofilament protein in transfected non-neuronal cells
Authors: Pleasure SJ , Lee VM , Nelson DL
Published: April 2018

We expressed the human midsized neurofilament subunit (NF-M) using genomic DNA in mouse L cells and showed that it is transcribed and translated into a protein capable of assembly into the cytoskeleton and of forming a filamentous network that colocalizes with the endogenous vimentin filaments. Moreover, human NF-M expressed in L cells is phosphorylated at sites within the multiphosphorylation repeat (MPR), i.e., the major sites of phosphorylation of NF-M in vivo. We also expressed a genomic construct lacking the MPR domain in the native molecule and showed that this MPR(-) protein also was expressed and formed a filamentous network despite diminished incorporation of radiolabeled phosphate. Two major conclusions emerged from the work described in this paper: human NF-M is translated, assembled, and phosphorylated at physiological sites without the need of any other specific neuronal proteins; phosphorylation sites other than the MPR are present within NF-M which may play a role in synthesis, assembly, and degradation of NF protein in humans.
Lineage, arrangement, and death of clonally related motoneurons in chick spinal cord
Authors: Leber SM , Breedlove SM , Sanes JR
Published: April 2018

We have used recombinant retroviruses as lineage markers to study the genealogy of motoneurons (MNs) in the chick spinal cord. We infected individual progenitors by injecting virions into the neural tube at stages 11–18, a few cell divisions before MNs are born. The descendants of infected cells were subsequently detected with a histochemical stain for beta-galactosidase (lacZ), the product of the retrovirally introduced gene. Clonally related, lacZ-positive cells formed clusters that were usually radial or planar in shape. The cells that comprised these clones were classified by morphology, size, and location. About 15% of the clones in the spinal cord contained MNs, and these were studied further. Multicellular clones that contained only MNs were infrequent. Instead, close relatives of MNs included a variety of other neurons, as well as glia and ependymal cells. Most non-MNs in these clones were found in the ventral and intermediate parts of the spinal cord. Neurons included interneurons and autonomic preganglionic neurons in the column of Terni. Labeled glia were found in both the gray and white matter and included astrocytes and cells tentatively identified as oligodendrocytes. Thus, even shortly before MNs are born, their progenitors are multipotential. Clonally related MNs were not restricted to a single motor pool. Some clones contained MNs in both the medial and lateral parts of the lateral motor column, which are known to innervate distinct groups of limb muscles. Furthermore, some clones contained MNs in the medial motor column (which innervate axial muscles) as well as in the lateral motor column. In contrast, the dispersal of clonally related MNs (and other neurons) was restricted in the rostrocaudal axis; most clones were less than one-quarter segment in length. Thus, MNs derived from a single progenitor are more likely to share rostrocaudal position than synaptic targets. To investigate the fate of clonally related MNs, we counted the number of MNs per clone at times before, during, and after the major period of MN death. The number of MNs per clone declined in precise parallel with the total number of MNs during this period, suggesting that neurons are eliminated without regard to their clone of origin. This result implies that the decision to live or die occurs on a cell-by-cell rather than a clone-by-clone basis.
Identification of location and timing of guidance cues in sympathetic preganglionic axons of the chick
Authors: Yip JW
Published: April 2018

The outgrowth of preganglionic axons into the sympathetic trunk of the chick occurs at stages 26–27 (5–5 1/2 d), beginning with C16 and T1 and followed by progressively more caudal spinal cord segments. The direction of preganglionic projections is segment specific, with T1 preganglionic neurons projecting predominantly in the rostral direction and T4 preganglionic neurons projecting predominantly in the caudal direction. These projections are correct from the outset. A previous study (Yip, 1987) has shown that removal of the neural crest, precursors of target ganglion cells, did not alter the trajectories of preganglionic axons, indicating that target cues are not required for the guidance of these axons. The present work extends that study and examines, through surgical manipulations of the developing embryo, the location and time of appearance of preganglionic axonal guidance cues. Spinal cord transplantation experiments show that segment-specific patterns of preganglionic outgrowth are determined by the local environment of axonal growth and not by the identity of preganglionic populations. In addition, spinal cord transplants of age-mismatched donors and hosts indicate that the timing of preganglionic outgrowth is also under local environmental control. These findings suggest that preganglionic axonal outgrowth is guided by local cues which may appear in a controlled spatiotemporal manner.
A study of the organellar Ca2(+)-transport ATPase isozymes in pig cerebellar Purkinje neurons
Authors: Plessers L , Eggermont JA , Wuytack F , Casteels R
Published: April 2018

Pig cerebellar Purkinje neurons express a high level of Ca2(+)- transport ATPases in their intracellular Ca2+ stores. This was shown at the mRNA level by Northern blotting and in situ hybridization and at the protein level by Western blotting and immunocytochemistry. The majority of the Ca2(+)-transport ATPases in these neurons belongs to the SERCA2b type (i.e., the Ca2(+)-pump isoform found in most nonmuscle cells). The SERCA2a (cardiac/slow-twitch skeletal/smooth muscle) Ca2(+)- pump isoform is expressed only at very low levels. The main Ca2(+)-pump messenger is 6.0 kilobases long and belongs to a class 4-type processing of SERCA2, which is exclusively confined to the cerebrum and cerebellum. Phospholamban, a regulator of the SERCA2 Ca2(+)-transport ATPase in cardiac/slow-twitch skeletal/smooth muscle, could not be detected in Purkinje neurons.
Channeling of developing rat corticospinal tract axons by myelin- associated neurite growth inhibitors
Authors: Schwab ME , Schnell L
Published: April 2018

CNS myelin contains 2 membrane proteins that are potent inhibitors of neurite growth (NI-35 and NI-250). Because myelin formation starts at different times in different regions and tracts of the CNS, this inhibitory property of myelin could serve boundary and guidance functions for late-growing fiber tracts. In the rat, the corticospinal tract (CST) grows into and down the spinal cord during the first 10 postnatal days, in close proximity to the sensory tracts fasciculus cuneatus and gracilis. Immunofluorescence for myelin constituents showed that, in the rostral half of the spinal cord, the myelinating tissue of these ascending tracts surrounds the growing, myelin-free CST in a channellike fashion. Elimination of oligodendrocytes by x- irradiation of the newborn rats, or application of antibody IN-1, which neutralizes the inhibitory substrate property of CNS myelin, resulted in significant anatomical aberration of CST fibers. In particular, the tract was larger in cross-section, and aberrant CST fibers and fascicles intermixed with the neighboring sensory ascending tracts. These results assign an important channeling and “guard-rail” function to the oligodendrocyte-associated neurite growth inhibitors for the developing CST in the rat spinal cord.
Calcium currents and graded synaptic transmission between heart interneurons of the leech
Authors: Angstadt JD , Calabrese RL
Published: April 2018

Synaptic transmission between reciprocally inhibitory heart interneurons (HN cells) of the medicinal leech was examined in the absence of Na-mediated action potentials. Under voltage clamp, depolarizing steps from a holding potential of -60 mV elicited 2 kinetically distinct components of inward current in the presynaptic HN cell: an early transient current that inactivates within 200 msec and a persistent current that only partially decays over several seconds. Both currents begin to activate near -60 mV. Steady-state inactivation occurs over the voltage range between -70 and -45 mV and is completely removed by 1–-2-sec hyperpolarizing voltage steps to -80 mV. The inward currents are carried by Ca2+, Ba2+, or Sr2+ ions, but not by Co2+, Mn2+, or Ni2+. These same inward currents underlie the burst-generating plateau potentials previously described in HN cells (Arbas and Calabrese, 1987a,b). With a presynaptic holding potential of -60 mV, the threshold for transmitter release is near -45 mV. Postsynaptic currents in the contralateral HN cell have a reversal potential near - 60 mV. The largest postsynaptic currents (300–400 pA) exhibit an initial peak response that is followed by a more slowly decaying component. The persistent component of Ca2+ current in the presynaptic neuron is strongly correlated with the prolonged component of the postsynaptic current, while the transient presynaptic Ca2+ current appears to correspond to the early peak of postsynaptic current. These data are consistent with the hypothesis that voltage-dependent calcium currents contribute to the oscillatory capability of reciprocally inhibitory HN cells by (1) generating the plateau potential that drives the burst of action potentials and (2) underlying the release of inhibitory transmitter onto the contralateral cell.
Development of serotonin-induced ion currents in identified embryonic Retzius cells from the medicinal leech (Hirudo medicinalis)
Authors: Lessmann V , Dietzel ID
Published: April 2018

Retzius cells and the cutaneous baroreceptive P-cells of adult medicinal leeches (Hirudo medicinalis) respond to 5-HT by Cl- as well as by monovalent cation conductances (Drapeau et al., 1989). However, chemical synaptic connections between Retzius cells (Liu and Nicholls, 1989) as well as Retzius cells and P-cells in culture (Drapeau and Sanchez-Armass, 1988) are mediated by postsynaptic Cl- currents in response to 5-HT release from the presynaptic cell. It was the aim of the present study to find out whether the response, which has been shown to be involved in synaptic transmission or whether the cation current, which has so far only been observed upon extrasynaptic 5-HT application, dominates during embryogenesis. Currents induced by transmitter application with a fast perfusion system were measured with a single-electrode voltage clamp and patch pipettes in whole-cell configuration. Control experiments, performed on Retzius cells from adult leeches after 1 d in culture, showed Cl- currents in response to 5-HT application in all cells investigated. Ninety percent of the cells tested showed additional 5-HT-elicited Na+, K+, or both currents. However, the conductance of cation currents amounted only to 10% of the Cl- currents. Embryonic Retzius cells were identified in dissociated cultures by their violet-blue fluorescence acquired by preincubation of intact ganglia or embryos in culture medium containing the autofluorescent 5-HT analog 5,7-dihydroxytryptamine. The first responses to 5-HT were measured at embryonic day 10 (E10), after voltage-activated Ca2+ and Na+ currents had already developed. 5-HT application induced exclusively Cl- currents until E16. This suggests that the receptor-channel complex, which also mediates synaptic responses, is expressed first in measurable quantities. Hence, there is currently no indication that extrasynaptic receptors dominate in immature cells in this preparation.
A mouse brain homolog of the Drosophila Shab K+ channel with conserved delayed-rectifier properties
Authors: Pak MD , Covarrubias M , Ratcliffe A , Salkoff L
Published: April 2018

We have cloned and expressed a mouse brain K+ channel that is the homolog of the Drosophila Shab K+ channel. Mouse and Drosophila Shab K+ channels (mShab and fShab, respectively) represent an instance of K+ channels and structurally related species that are both functionally and structurally conserved; most kinetic, voltage-sensitive, and pharmacological properties are similar for the 2 channels. The greatest functional difference between the currents is recovery from inactivation, which is several times slower for mShab than for fShab currents. In addition to conserved structure, the mShab polypeptide has an unusually long nonconserved region at the carboxyl end of the protein. Truncation of 293 residues from the carboxyl end produced no noticeable change in voltage-sensitive, kinetic, or pharmacological properties. Thus, the measured functional properties of mShab are determined by the remaining 564 residues, most of which are conserved. The mShab and fShab channels are naturally occurring structural variants having substitutions in conserved portions that appear relatively neutral with respect to all measured properties except for, possibly, the rate of recovery from inactivation. The mShab current closely resembles a native delayed-rectifier-type potassium current, IK, in hippocampal neurons.
Inhibition of glucocorticoid secretion by the hippocampal formation in the primate
Authors: Sapolsky RM , Zola-Morgan S , Squire LR
Published: April 2018

Inhibition of the adrenocortical axis by glucocorticoids (GCs) occurs at both hypothalamic and suprahypothalamic sites. In the rat, the hippocampus has been shown to be an essential suprahypothalamic site. The present study shows that the hippocampal system serves a similar role in the nonhuman primate. Bilateral lesions that included the hippocampal formation and the parahippocampal cortex; the hippocampal formation, parahippocampal cortex, and the amygdala; or the fornix all produced GC hypersecretion in cynomolgus monkeys. The hypersecretion occurred throughout the day. Moreover, these lesions were also associated with dexamethasone resistance (i.e., GC hypersecretion following administration of the synthetic GC dexamethasone). The hypersecretion could not be attributed to acute surgical trauma, because neither circumscribed lesions of the amygdala nor conjoint lesions of the perirhinal and parahippocampal cortex produced adrenocortical abnormalities. Finally, in agreement with data derived from the rat, the GC hypersecretion following hippocampal lesions was transient. Secretory activity returned to normal levels by 6-15 months in all operated groups. Thus, the primate hippocampal system appears to share some neuroendocrine functions with the rodent.
Evaluation of projection patterns in the primary olfactory system of rainbow trout
Authors: Riddle DR , Oakley B
Published: April 2018

Topographic projections are important for coding sensory information in the visual, auditory, and somatosensory systems but are of uncertain importance in the coding of olfactory information. We searched for topographic projections between olfactory receptor cells and the olfactory bulb of the rainbow trout Oncorhynchus mykiss. Anterograde axonal tracing with HRP revealed that the olfactory axons arising from discrete regions of the olfactory epithelium travel together within the olfactory nerve. The abrupt resorting and redistribution of these axons at the interface between the olfactory nerve and olfactory bulb imply that local cues control and organize axonal projections. The sites of termination of HRP-labeled axons in the glomerular layer could not be predicted from the location of their cell bodies in the periphery. Retrograde tracing with fluorescently labeled latex beads, injected into glomerular subregions as small as 1% of the total glomerular volume, labeled receptor cells dispersed throughout the olfactory epithelium. The distributions of labeled receptor cells were uncorrelated with the bulbar injection sites. Double-labeling experiments revealed that even widely separated sites in the glomerular layer receive axons from comingled populations of receptor cells. Hence, the evidence indicates that the spatial arrangement of olfactory receptor cells in the epithelium is not preserved in the termination of their axons in the olfactory bulb. We conclude that the primary olfactory in trout lacks point-to-point or regionally topographic organization and that the entire extent of the olfactory epithelium contributes axons to each region of the glomerular layer.