bluuebunny/crossref_metadata_2025 huggingface dataset:
Lesions of the periaqueductal gray and rostral ventromedial medulla disrupt antinociceptive but not cardiovascular aversive conditional responses
Authors: Helmstetter FJ , Tershner SA
Published: April 2018

The presentation of an auditory stimulus that signals a noxious event such as foot shock results in the simultaneous expression of multiple aversive conditional responses (CRs), which include a transient elevation of arterial blood pressure (ABP) and an opioid-mediated form of hypoalgesia. Recent evidence suggests that the neural circuits responsible for the expression of these two aversive responses may overlap. In the present study, rats were trained using a Pavlovian fear conditioning paradigm in which white noise was repeatedly paired with shock. After training, groups of animals received electrolytic lesions centered in the dorsal or ventral periaqueductal gray (PAG) or in the medial or lateral rostral medulla. In sham-lesioned animals that were given paired presentations of noise and shock, subsequent presentation of the auditory stimulus caused a significant transient elevation of ABP and time-dependent inhibition of the tail flick reflex evoked by radiant heat. Lesions of either the dorsal or the ventral PAG blocked the antinociceptive CR but did not significantly affect ABP responses. Lesions of the ventromedial, but not the lateral, rostral medulla blocked hypoalgesia. Rostral medullary lesions did not reliably affect stimulus-evoked cardiovascular responses or baseline ABP. These results indicate that antinociceptive and cardiovascular conditional responses are anatomically dissociable and support our proposal that conditional hypoalgesia is mediated by a serial neural circuit that includes the amygdala, PAG, and rostral ventromedial medulla.
Modulation of Ca2+ channels by PTX-sensitive G-proteins is blocked by N- ethylmaleimide in rat sympathetic neurons
Authors: Shapiro MS , Wollmuth LP , Hille B
Published: April 2018

The actions of N-ethylmaleimide (NEM), a sulfhydryl alkylating agent, on G-protein-mediated inhibition of N-type Ca2+ channels in adult rat superior cervical ganglion (SCG) neurons were studied using whole-cell voltage clamp. In SCG neurons, inhibition of ICa occurs by at least three separable pathways: one pertussis toxin (PTX) sensitive and voltage dependent, and two PTX insensitive and voltage independent. NEM blocked PTX-sensitive inhibition nearly completely, with only small effects on PTX-insensitive inhibition. Somatostatin inhibition is completely PTX sensitive and was wholly blocked by a 120 sec exposure to 50 microM NEM, with shorter exposure times producing a less complete block. Inhibition of ICa by norepinephrine (NE) is approximately half PTX sensitive and was also approximately half NEM sensitive. One component of muscarinic inhibition is PTX insensitive, voltage independent, and mediated by a diffusible cytoplasmic messenger; this pathway was largely spared by NEM treatment. Another pathway is also PTX insensitive and voltage independent, used by substance P, and was also largely NEM insensitive. Hence, in SCG neurons, NEM selectively inactivates PTX-sensitive G-proteins. We also find evidence that the PTX-insensitive action of NE is distinct from the other PTX-insensitive pathways, and therefore assign it to a fourth signaling pathway.
Behavioral associations of neuronal activity in the ventral tegmental area of the rat
Authors: Kosobud AE , Harris GC , Chapin JK
Published: April 2018

The ventral tegmental area (VTA) is a central element in a system that mediates the reinforcing properties of natural stimuli (such as food), brain stimulation, and drugs of abuse. Although considerable effort has been applied to understanding how drugs of abuse influence this system, relatively little work has examined its function during conditioned reinforcement tasks in awake, behaving animals. In the present studies, bundles of four to eight microwire electrodes were chronically implanted in the VTA or prefrontal cortex (PFC) of male Wistar rats. Following recovery from surgery, simultaneous recordings from multiple single neurons and unit clusters were obtained in rats pressing a lever for a sucrose solution on a fixed-ratio schedule of reinforcement. Consistent with the hypothesis that these neurons encode information related to motivation, most of the neurons in both VTA and PFC showed significant modulation of firing rate associated with one or more events occurring within the response/reinforcement cycle. These events included lever presses, onset and end of a tone signaling sucrose delivery, and onset and end of sucrose consumption. Significant decreases in firing rate were observed, coincident with onset of the tone and sucrose delivery, or with consumption. These decreases were sustained through the end of sucrose consumption. A number of neurons also discharged bursts of activity associated with individual lever presses. These findings provide a clear demonstration that VTA neuronal activity is modulated during motivated behavior. Similar information about events within the ongoing response/reinforcement cycle appears to be distributed through many neurons within the VTA, and may be mirrored in target structures such as PFC.
Receptive field analysis and orientation selectivity of postsynaptic potentials of simple cells in cat visual cortex
Authors: Pei X , Vidyasagar TR , Volgushev M , Creutzfeldt OD
Published: April 2018

Postsynaptic potentials (PSPs) were recorded from cat striate cortical cells by the whole-cell in vivo recording technique using patch-clamp electrodes. EPSPs and IPSPs evoked by flashing bars on the receptive field at different positions and orientations revealed the spatial structure of the excitatory and inhibitory inputs. The elongation of the excitatory input field (length:width ratio) was found to be minimal (mean ratio of 1.7) and much lower than those reported for spike discharges. Two-dimensional receptive field response profiles of early PSPs were recorded by flashing a small spot of light over a square matrix covering the receptive field. These recordings also showed only mild degrees of elongations of the receptive field. Such elongations could be the result of either an excitatory input from the geniculate that is already biased for orientation or an excitatory convergence from a limited number of LGN fields arranged in a row. In most first- order cells, we found that inhibition was contributing significantly to orientation selectivity. Often prominent IPSPs could be evoked by stimuli of nonoptimum orientations. Presence of inhibition could also be inferred by the way that EPSPs were sharply cut off by inhibition. When the amplitude of an EPSP was measured at different latencies after its onset, the EPSP was found to be very broadly tuned to orientation at the beginning, but showing increasing orientation selectivity with time. It is proposed that this progressive development of orientation selectivity is due to (1) inhibitory inputs arriving after the first wave of excitation, (2) intracortical excitatory inputs from other cells tuned to similar orientations, and (3) voltage-sensitive mechanisms such as NMDA channels.
Activation of postsynaptic metabotropic glutamate receptors by trans- ACPD hyperpolarizes neurons of the basolateral amygdala
Authors: Rainnie DG , Holmes KH , Shinnick-Gallagher P
Published: April 2018

Glutamate has traditionally been regarded as an excitatory neurotransmitter. Synaptic activation of ionotropic glutamate receptors mediates fast EPSPs in the CNS. Moreover, activation of metabotropic glutamate receptors (mGluRs), which are coupled to second messenger effector systems via GTP-binding proteins (G-proteins), results in the expression of slow EPSPs. We have now examined the response of basolateral amygdala (BLA) neurons to activation of postsynaptic mGluRs. In approximately 78% of BLA neurons examined, activation of postsynaptic mGluRs results in membrane hyperpolarization and an associated decrease in membrane input resistance or a hyperpolarization followed by a depolarization associated with an increase in input resistance. The purpose of this study was to address the mechanisms underlying the membrane hyperpolarization. Here, we report that the ACPD-induced hyperpolarization is insensitive to TTX, is dependent on extracellular K+ concentrations, and has a reversal potential (-84 mV) close to that estimated from the Nernst equation for an increase in a K+ conductance. In addition, the ACPD response is resistant to (1) intracellular chloride loading, (2) the GABAB receptor antagonist CGP55845A, (3) the ACh receptor antagonist atropine, and (4) the ionotropic glutamate receptor antagonists CNQX and APV. These data suggest that the hyperpolarization results from a direct activation of postsynaptic mGluRs on neurons of the BLA. Furthermore, we performed studies that suggest that the hyperpolarization is G-protein mediated and results from activation of a TEA-sensitive, calcium-dependent potassium conductance. The sensitivity of this conductance to thapsigargin further suggests that this response requires the release of calcium from intracellular stores. In summary, these data suggest a role for glutamate as an inhibitory transmitter in the BLA during periods of metabotropic glutamate receptor activation. In nuclei such as the BLA that are exquisitely sensitive to seizure induction, an inhibitory response to glutamate may act to delay the onset of epileptogenesis.
On the directional firing properties of hippocampal place cells
Authors: Muller RU , Bostock E , Taube JS , Kubie JL
Published: April 2018

Using a two-spot tracking system that allowed measurements of the direction of a rat's head in the environment as well as the position of the rat's head, we investigated whether hippocampal place cells show true direction-specific as well as location-specific firing. Significant modulations of firing rate by head direction were seen for most cells while rats chased food pellets in a cylindrical apparatus. It was possible, however, to account quantitatively for directional modulation with a simple scheme that we refer to as the “distributive hypothesis.” This hypothesis assumes that firing is ideally location specific, and that all directional firing modulations are due to differences in the time that the rat spends in different portions of the firing field of the place cell in different head direction sectors. When the distributive hypothesis is put into numeric form, the directional firing profiles that it predicts are extremely similar to the observed directional firing profiles, strongly suggesting that there is no intrinsic directional specificity of place cell firing in the cylinder. Additional recordings made while rats ran on an eight-arm maze reveal that many firing fields on the arms are polarized; the cell discharges more rapidly when the rat runs in one direction than the other on the maze. This result provides an independent confirmation of the findings of McNaughton et al. (1983). For fields that appear to be polarized by inspecting firing rate maps of the raw data, the magnitude of directional firing variations is greater than predicted by the distributive hypothesis. By comparison with postsubicular head direction cells, it is shown that the distributive prediction of weaker- than-observed directional firing is expected if there is a true directional firing component. A major conclusion reached from recording in both environments is that the directional firing properties of hippocampal place cells are variable and not fixed; this is true of individual units as well as of the population.
Differential effects of morphine on noxious stimulus-evoked fos-like immunoreactivity in subpopulations of spinoparabrachial neurons
Authors: Jasmin L , Wang H , Tarczy-Hornoch K , Levine JD , Basbaum AI
Published: April 2018

In previous studies we reported that although morphine dose dependently inhibits noxious stimulus-evoked expression of the c-fos proto-oncogene in the rat spinal cord, morphine was without effect in certain populations of presumed nociresponsive neurons, even under conditions of complete behavioral analgesia. To determine whether the neurons that continue to express the c-fos gene include projection neurons, we evaluated the effect of morphine on noxious stimulus-evoked c-fos expression in spinoparabrachial neurons retrogradely labeled with Fluoro-gold. In the formalin test, we found that morphine analgesia was associated with a significant reduction in the number of Fos-like- immunoreactive spinoparabrachial projection neurons in the lateral reticulated area of the neck of the dorsal horn. Morphine, however, did not reduce the number of Fos-like-immunoreactive spinoparabrachial projection neurons either in the superficial dorsal horn or in the area around the central canal. These results indicate that under conditions of morphine analgesia two distinct populations of spinoparabrachial neurons can be recognized on the basis of their expression of the c-fos gene in response to noxious stimulation. Since the expression of the c- fos gene has been correlated with neuronal activity, these data suggest that activity, and central transmission of nociceptive information, persists in certain nociresponsive projection neurons during morphine analgesia. Alternatively, if activity has, in fact, been blocked in these neurons, our results indicate that injury can produce significant molecular changes in neurons even though the neuronal activity and pain associated with the injury is blocked by morphine.
Opioids modulate stress-induced proenkephalin gene expression in the hypothalamus of transgenic mice: a model of endogenous opioid gene regulation by exogenous opioids
Authors: Borsook D , Falkowski O , Rosen H , Comb M , Hyman SE
Published: April 2018

Stressful stimuli strongly induce proenkephalin gene expression within the paraventricular nucleus (PVN) of the hypothalamus. A human proenkephalin-beta-galactosidase fusion gene has previously been shown to give correct phenotypic expression and appropriate stress regulation within the hypothalamus of transgenic mice; this model provides high sensitivity, cellular resolution, and ready quantification of levels of proenkephalin gene expression. Here we describe use of this transgenic model to study modulation of stress-regulated gene expression in the PVN by opiates. Acute or subacute morphine administration prior to a hypertonic saline stress produced marked superinduction of transgene expression compared with hypertonic saline stress alone. In contrast, chronic morphine administration decreased basal expression of the transgene, and inhibited stress-induced expression of the transgene. The endogenous proenkephalin mRNA was induced in parallel with the transgene as demonstrated by in situ hybridization; the immediate-early gene c-fos was also regulated in parallel with the transgene. These data suggest that acute or subacute morphine administration sensitizes proenkephalin neurons within the PVN and other regions of the hypothalamus to stress and that chronic morphine administration desensitizes this response. Because the molecular mechanisms regulating the expression of the transgene are well understood, this model provides a useful tool for investigating cellular and molecular effects of opioids on the hypothalamus.
A Drosophila receptor tyrosine phosphatase expressed in the embryonic CNS and larval optic lobes is a member of the set of proteins bearing the "HRP" carbohydrate epitope
Authors: Desai CJ , Popova E , Zinn K
Published: April 2018

Recent studies have defined several cell surface glycoproteins expressed in the developing nervous system of insect embryos that may be involved in axon outgrowth and guidance processes. These glycoproteins include the fasciclins and a group of receptor-linked protein tyrosine phosphatases (R-PTPs). In embryos, the fasciclins are localized to axonal subsets, while the R-PTPs appear to be expressed on most or all CNS axons. To identify other neuronal cell surface glycoproteins in the Drosophila embryo, we have taken a biochemical approach. This is based on the observation that antisera against horseradish peroxidase (HRP) recognize a carbohydrate epitope that is selectively expressed in the insect nervous system. A large number of neuronal glycoproteins (denoted “HRP proteins”) apparently bear the HRP carbohydrate epitope. We have used polyclonal anti-HRP antibodies to purify these proteins from Drosophila embryos, and have obtained protein sequences from seven HRP protein bands. These data define three major HRP proteins as neurotactin, fasciclin I, and an R-PTP, DPTP69D. Western blotting data suggest that fasciclin II, neuroglian, DPTP10D, and DPTP99A are also HRP proteins. We show that DPTP69D, like the previously characterized R-PTPs, is localized to CNS axons in the embryo. In third instar larvae, DPTP69D expression is restricted to subsets of neuronal processes in the brain, ventral nerve cord, and eye disk. In the optic lobes, DPTP69D is localized to the neuropils of the lamina and medulla, and to an array of parallel thick bundles that may be the transmedullary fibers of the developing lobula complex.
Functional mapping of horizontal connections in developing ferret visual cortex: experiments and modeling
Authors: Weliky M , Katz LC
Published: April 2018

In cat striate cortex, patchy horizontal axonal projections link columns of similar orientation specificity. To assess the physiological correlates of such clustered projections, a new multisite stimulation technique was used to functionally map the pattern of horizontal synaptic inputs onto single layer 2/3 cells within tangential slices of developing ferret visual cortex. Twenty-four separate sets of horizontal fibers were stimulated within a 1200 microns strip of cortex, while evoked synaptic responses were recorded using whole-cell patch methods. For most cells, input maps demonstrated the presence of clustered horizontal connections in which multiple strong and weak synaptic responses were alternately evoked across the stimulated cortical region. Recordings from up to nine cells in a single slice revealed that patterns of synaptic input were closely correlated for cells in close proximity, and that this correlation decreased with distance, with no correlation at distances greater than 500 microns. To determine whether these physiological results were consistent with the known anatomical linkage of iso-orientation columns by clustered horizontal connections, mathematical analysis and computer simulations were performed upon orientation tuning maps obtained from optical imaging of activity-dependent intrinsic signals in mature ferret visual cortex. Optical imaging revealed an organization of iso-orientation domains consisting of broad regions of cortex across which orientation preference smoothly varied, together with “orientation centers” around which orientation preference was arranged in a pinwheel manner. The distribution of synaptic connections between different cortical sites was simulated by a model of functionally linked iso- orientation columns. Simulated synaptic input maps, generated by the same stimulation and recording arrangements used in our experimental protocol, accurately reproduced the observed patterns of clustered inputs onto experimentally recorded cells. These results indicate that even at the time of eye opening, prior to extensive visual experience, most cells receive patterns of synaptic inputs consistent with a clustered organization of horizontal connections that functionally link iso-orientation columns.
Immunocytochemical localization and biological activity of 3 beta- hydroxysteroid dehydrogenase in the central nervous system of the frog
Authors: Mensah-Nyagan AG , Feuilloley M , Dupont E , Do-Rego JL , Leboulenger F , Pelletier G , Vaudry H
Published: April 2018

The enzyme 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase (3 beta-HSD) catalyzes biosynthesis of progesterone (P) and all precursors of glucocorticoids, mineralocorticoids, androgens, and estrogens. Despite the broad interest raised by neurosteroids, the cellular localization of 3 beta-HSD has never been investigated in the brain. We took advantage of the availability of an antiserum raised against human placental 3 beta-HSD to determine the distribution of 3 beta-HSD-immunoreactive structures in the brain of the frog Rana ridibunda by the indirect immunofluorescence technique. Three populations of 3 beta-HSD-immunoreactive cell bodies were observed in the hypothalamus, namely, in the rostral region of the preoptic nucleus, the dorsal infundibular nucleus, and the dorsal part of the ventral infundibular nucleus. A dense network of 3 beta-HSD- immunoreactive nerve fibers was visualized in the dorsal area of the diencephalon, that is, in the lateral neuropil, the corpus geniculatus lateralis, and the nucleus posterolateralis thalami. Reversed-phase HPLC analysis of frog hypothalamic extracts combined with RIA detection showed the presence of substantial amounts of immunoreactive steroids coeluting with P and 17-hydroxyprogesterone (17OH-P). The synthesis of delta 4–3-keto-steroids in the frog hypothalamus was investigated using the pulse-chase technique with 3H-pregnenolone (3H-delta 5P) as a precursor. The formation of five tritiated metabolites of 3H-delta 5P was observed, one of which coeluted with 17OH-P. Conversion of 3H-delta 5P into this radioactive metabolite was significantly reduced by trilostane, a specific inhibitor of 3 beta-HSD. Immunodetection of newly synthesized steroids in HPLC fractions of hypothalamic extracts, using 17OH-P antibodies, revealed the existence of an immunoreactive steroid that exhibited the same retention time as synthetic 17OH-P. The present study provides the first immunocytochemical mapping of 3 beta- HSD, a key enzyme of the steroid biosynthetic pathway, in the CNS of a vertebrate. The data also demonstrate for the first time biosynthesis of neurosteroids in the brain of a nonmammalian vertebrate.
The T alpha 1 alpha-tubulin promoter specifies gene expression as a function of neuronal growth and regeneration in transgenic mice
Authors: Gloster A , Wu W , Speelman A , Weiss S , Causing C , Pozniak C , Reynolds B , Chang E , Toma JG , Miller FD
Published: April 2018

We have previously demonstrated that one member of the alpha-tubulin multigene family, termed T alpha 1 in rats, is regulated as a function of neuronal growth and regeneration. To elucidate the molecular mechanisms responsible for coupling gene expression to morphological differentiation, we have isolated the T alpha 1 gene, have fused 1.1 kb of the 5′ flanking region to a nuclear lacZ reporter gene, and have generated transgenic mice. Analysis of these transgenic mice demonstrated that marker gene expression was specific to the CNS and PNS, with expression in vivo at embryonic day 13.5 being similar to expression of the endogenous gene. Moreover, the induction of transgene expression was correlated temporally with neuronal commitment in developing neural crest-derived peripheral neurons and in the developing retina. Immunocytochemical analysis of mixed primary embryonic brain cultures confirmed that transgene expression was specific to neurons, with the majority of neurons, but not astrocytes or oligodendrocytes, expressing beta-galactosidase. Transgene expression in vivo was maintained in developing neurons until early in postnatal life, subsequent to which its expression decreased coincident with neuronal maturation. The transgene was then reinduced in regenerating facial motoneurons following unilateral axotomy of the facial nerve. Thus, 1.1 kb of 5′ flanking sequence from the T alpha 1 gene contains the sequence elements responsible for specifying gene expression to embryonic neurons and for subsequently regulating gene expression in both developing and mature neurons as a function of morphological growth.
Transparent motion perception as detection of unbalanced motion signals. III. Modeling
Authors: Qian N , Andersen RA , Adelson EH
Published: April 2018

In the preceding two companion articles we studied the conditions under which transparent motion perception occurs through psychophysical experiments, and investigated the underlining neural mechanisms through physiological recordings. The main finding of our perceptual experiments was that whenever a display has finely balanced motion signals in all local areas, it is perceptually nontransparent, and that transparent displays always contain motion signals in different directions that are either spatially unbalanced, or unbalanced in their disparity or spatial frequency contents. In the physiological experiments, we found two stages in the processing of transparent stimuli. The first stage is located primarily in area V1. At this stage motion measurements are made and V1 cells respond well to both the balanced, nontransparent stimuli and the unbalanced, perceptually transparent stimuli. The second stage is located primarily in area MT. MT cells show strong suppression between opposite directions of motion. The suppression for the unbalanced, transparent stimuli is significantly less than that for the balanced, nontransparent stimuli. Therefore, the activity in the second, MT stage correlates better with the perception of motion transparency than the first, V1 stage, which does not distinguish reliably between transparent and nontransparent motion. The above experiments suggest a two-stage model of motion perception with a motion measurement stage in V1 and an opponent- direction suppression stage in area MT. In this article we explicitly test this model through analysis and computer simulations, and compare the response of the model to the perceptual and physiological results using the same balanced and unbalanced stimuli we used in the experiments. In the first stage of the computational model, motion energies in different spatial frequency and disparity ranges are extracted from each local region. Similar to V1, this stage does not distinguish between the balanced and unbalanced stimuli. In the subsequent stage motion energies of opposite directions but with same spatial frequency and disparity contents suppress each other using subtractive or divisive inhibition. This stage responds significantly better to the transparent stimuli than to the nontransparent ones, in agreement with MT activity.
Bidirectional promoter of human monoamine oxidase A (MAO A) controlled by transcription factor Sp1
Authors: Zhu QS , Chen K , Shih JC
Published: April 2018

The core promoter region of human monoamine oxidase (MAO) A has been identified in the two 90 bp repeat sequences, which can be further divided into four imperfect tandem repeats, each containing an Sp 1 binding site in the reversed orientation. Gel retardation and DNase 1 footprinting assays identified Sp 1 to be the major transcription factor binding to MAO A core promoter. In addition, positive association has been observed between cellular Sp1 concentration and MAO A promoter or catalytic activity, indicating that Sp1 is a controlling factor for human MAO A expression. DNA fragments from MAO A core promoter exhibit promoter activity in both orientations in a transient transfection assay, using human growth hormone as the reporter gene. A DNA probe isolated from upstream of the core promoter detected positive signals in a Northern analysis, suggesting that the reverse promoter activity may endogenously transcribe a new gene located upstream of MAO A.
Inhibition of nitric oxide synthase does not impair spatial learning
Authors: Bannerman DM , Chapman PF , Kelly PA , Butcher SP , Morris RG
Published: April 2018

Nitric oxide (NO), a putative intercellular messenger in the CNS, may be involved in certain forms of synaptic plasticity and learning. This article reports a series of experiments investigating the effects of N omega-nitro-L-arginine methyl ester (L-NAME) upon various forms of learning and memory in the watermaze. L-NAME (75 mg/kg, i.p., sufficient to bring about > 90% inhibition of NO synthesis in brain) produced an apparent impairment in spatial learning when given to naive rats during acquisition (3 d, six training trials per day). This impairment was dose related, stereoselective, and attenuated by coadministration of L-arginine. A second study showed that L-NAME did not affect the retention of a previously learned spatial task. In addition, in a visual discrimination task, the rate at which criterion levels of performance were reached was unaffected by L-NAME. Thus, inhibition of NO synthase may cause a selective impairment of spatial learning without effect upon retention. However, analysis of the early training trials of the visual discrimination task revealed significantly elevated escape latencies in the L-NAME-treated rats, suggesting that inhibition of NO synthase may have more general effects. As normal rats learn the spatial task very rapidly, the possibility arises that the apparent deficit in learning is due to a disruption of some process other than learning per se. A further series of experiments investigated this possibility. L-NAME was found not to impair the learning of a new platform position in the same spatial environment. Surprisingly, L-NAME also had no effect on spatial learning in a second watermaze located in a novel spatial environment by rats well practiced with all aspects of watermaze training. Finally, L-NAME had no effect on spatial learning in naive rats trained with just one trial per day. Thus, systemic injection of an NO synthase inhibitor impairs behavioral performance in two tasks during their initial acquisition, but the basis of this functional disruption is unlikely to be due to any direct effect upon the mechanisms of spatial learning.
Task-dependent effects of intra-amygdala morphine injections: attenuation by intra-amygdala glucose injections
Authors: Ragozzino ME , Gold PE
Published: April 2018

Intraseptal injections of morphine impair learning and memory in rats, and these impairments are reversed by intraseptal injections of glucose. With evidence that injections of morphine into the amygdala also impair memory for some tasks, the present experiment determined whether (1) intra-amygdala morphine injections impair performance in inhibitory avoidance and spontaneous alternation tasks, and (2) intra- amygdala glucose injections attenuate the effects of intra-amygdala morphine injections. Rats receiving bilateral injections of morphine (4.0 nmol) into the amygdala, 30 min prior to training in inhibitory avoidance, had retention latencies significantly lower than those of unoperated and CSF controls when tested 24 hr later. Bilateral morphine injections (4.0 or 8.0 nmol) 30 min prior to testing in a spontaneous alternation task did not alter performance. The morphine-induced impairment observed in inhibitory avoidance was not due to diffusion up the cannulas, altered sensitivity to shock, or seizure activity. A glucose dose of 16.67 nmol, but not 8.33 nmol, injected into the amygdala attenuated the morphine-induced deficit in inhibitory avoidance. Rats receiving CSF into the amygdala exhibited decreased retention latencies in inhibitory avoidance compared to those of unoperated controls. This decrease was not attenuated by glucose at doses of 8.33 and 16.67 nmol. Therefore, these findings suggest that the amygdala is another brain region in which glucose affects brain functions, possibly by interacting with the opioid system and/or other neurotransmitter systems.
Calcium-activated release of nitric oxide and cellular distribution of nitric oxide-synthesizing neurons in the nervous system of the locust
Authors: Muller U , Bicker G
Published: April 2018

Nitric oxide (NO) is generated by a Ca2+/calmodulin-activated NO synthase and diffuses as a short-lived transcellular messenger through the plasma membrane. This study investigates the neurochemistry and anatomical distribution of NO-releasing cells in the CNS of the locust. Ca2+/calmodulin-activated NO synthase is responsible for fixation- sensitive NADPH diaphorase (NADPHd) activity in cell homogenates of the nervous system. Therefore, neurons expressing NO synthase were detected by NADPHd histochemistry performed in whole-mounts. The anatomical screening revealed fewer than 1% NADPHd-positive cells in the ventral nerve cord, some of which were single potentially identifiable neurons, and groups of cell bodies in several regions of the cerebral ganglion. A prominent feature of the histochemical survey in the cerebral ganglion is a group of 45 intensely stained cells innervating the olfactory neuropil of the antennal lobe. A basic requirement for identifying NO as a messenger molecule is the Ca(2+)-dependent release during nerve cell depolarization. With a sensitive photometric assay we demonstrated that dissociated cells from brain areas rich in NADPHd- positive neurons release NO after stimulation by agents elevating cytoplasmic Ca2+ levels and by the excitatory neurotransmitter acetylcholine. The combined anatomical and biochemical experiments therefore provide firm evidence that NO is a messenger molecule released in the CNS of the locust. Since locust neurons can be readily grown in primary culture, NO-induced elevations of CGMP levels and other signal transduction mechanisms in target cells will also be amenable to a cellular analysis.
Long-term delayed vascularization of human neural transplants to the rat brain
Authors: Geny C , Naimi-Sadaoui S , Jeny R , Belkadi AM , Juliano SL , Peschanski M
Published: April 2018

Human neural transplants are being developed to treat Parkinson's disease. Previous characterization of human transplants focused on neuronal development, while little is known of the interaction between the transplant and its environment, among which blood is of prime importance. We evaluated here the formation of blood vessels in human neural xenografts placed into the brain of rats immunosuppressed with cyclosporin A. Using capillary wall markers, we found that human transplants remain virtually nonvascularized for more than 1 month. Angiogenesis takes place very slowly and the density of blood vessels is still quite poor after 3 months, the fine structure of these capillaries, when they form, is apparently normal. Functional studies indicate that the vascular network formed in the transplant allows blood circulation and exhibits a working barrier to macromolecules. Glucose uptake and consumption and cytochrome oxidase activity are almost undetectable up to 3 months after grafting. These results demonstrate that vascularization is much delayed in human xenografts into the rat brain. This delay is likely to be dependent on the maturation of the transplanted tissue. A dedifferentiation of human endothelial cells cotransplanted with neural cells occurs since histochemical and immunocytochemical markers revealing endothelial cells in the human fetus are not present up to 1 month in the transplant. The origin of this phenomenon is a matter of speculation. How neural cells survive and mature in such conditions are issues of prime interest for the future of human neural grafting.
Cyclic AMP and synaptic activity-dependent phosphorylation of AMPA- preferring glutamate receptors
Authors: Blackstone C , Murphy TH , Moss SJ , Baraban JM , Huganir RL
Published: April 2018

Several studies have suggested that the function of glutamate receptor channels can be regulated by protein phosphorylation. Furthermore, a basal level of phosphorylation may be necessary to maintain receptor function. Little is known, however, about the phosphorylation state of glutamate receptor channels in neurons and how it is regulated by synaptic activity. In this study, we have investigated the phosphorylation of the AMPA-preferring glutamate receptor subunit GluR1 in cortical neurons in primary culture. These neurons elaborate extensive processes, form functional synapses, and exhibit spontaneous 4–8 sec bursts of synaptic activity every 15–20 sec. In cultures in which this synaptic activity was suppressed by tetrodotoxin and MK-801, the GluR1 protein was phosphorylated on serine residues within a single tryptic phosphopeptide, as determined by phosphoamino acid analysis and phosphopeptide mapping. This same peptide was basally phosphorylated in recombinant GluR1 receptors transiently expressed in human embryonal kidney 293 cells. Treatment of these synaptically inactive cortical neurons with the adenylyl cyclase activator forskolin resulted in a robust increase in phosphorylation on serine residues on a phosphopeptide distinct from the basally phosphorylated peptide. Again, this same phosphopeptide was observed in recombinant GluR1 receptors isolated from 293 cells coexpressing the catalytic subunit of cAMP- dependent protein kinase. Spontaneous synaptic activity in cultures of cortical neurons resulted in a consistent, rapid (within 10–30 sec) increase in phosphorylation on serine and threonine residues. Interestingly, these phosphopeptides were also phosphorylated when neurons from inactive cultures were stimulated with phorbol esters, which activate protein kinase C. These results indicate that AMPA receptors containing the GluR1 subunit may be regulated by extracellular signals working through the cAMP second messenger system as well as by synaptic activity, possibly acting through protein kinase C. Such regulation by protein phosphorylation may be involved in short- term changes in synaptic efficacy thought to involve the functional modulation of AMPA receptors.
Effect of serotonin depletion on vibrissa-related patterns of thalamic afferents in the rat's somatosensory cortex
Authors: Bennett-Clarke CA , Leslie MJ , Lane RD , Rhoades RW
Published: April 2018

To investigate the role of 5-HT in the development of the somatosensory cortex, this amine was depleted in newborn (P-0) rats with a single subcutaneous injection of the toxin 5,7-dihydroxytryptamine (5,7-DHT) and thalamocortical organization was assayed by application of the carbocyanine dye Di-I to the thalamocortical radiations or ventrobasal thalamus, or by staining cortical sections for AChE or cytochrome oxidase (CO). High-performance liquid chromatographic analysis of cortices from animals killed on P-6 or P > 60 demonstrated that 5,7-DHT treatment resulted in 85.04 +/- 12.6% and 72.5 +/- 1.5% reductions in cortical 5-HT, respectively. Alternate cortices from the brains of animals killed on P-6 processed for 5-HT immunoreactivity demonstrated a complete absence of the vibrissa-related pattern of immunoreactivity and only a small number of coarse immunoreactive axons. The 85% depletion of 5-HT did not alter the somatotopic organization of thalamocortical afferents in animals killed on P-6 or P > 60, but it did cause 30.5 +/- 7.3% and 19.1 +/- 3.7% reductions in the cross- sectional areas of the patches of thalamocortical afferents corresponding to the long mystacial vibrissae (p < 0.05). These reductions were not associated with significant reductions in either brain or cortical weight or with decreases in the dimensions of the thalamic representation of the vibrissa follicles. These results indicate that 5-HT plays a significant role in the development of the thalamic innervation of the primary somatosensory cortex.
Inhibition of neurite growth by the NG2 chondroitin sulfate proteoglycan
Authors: Dou CL , Levine JM
Published: April 2018

The chondroitin sulfate proteoglycans (CSPGs) have been implicated as both positive and negative modulators of axonal growth; however, the functional properties of only a few specific CSPGs have been investigated. Here we demonstrate that NG2, an integral membrane CSPG expressed on the surfaces of glial progenitor cells, inhibits neurite growth from neonatal rat cerebellar granule neurons when presented to the cells as a component of the substrate. Growth inhibition occurred when NG2 was mixed with either laminin or L1, two potent promoters of axonal extension. Moreover, when given a choice between surfaces coated with NG2 and laminin or L1, the axons of the cerebellar neurons extended preferentially on laminin or L1 and avoided areas of the substrate containing NG2. The NG2 proteoglycan inhibited neurite growth after digestion with chondroitinase ABC, demonstrating that the inhibitory activity is a property of the core protein and not the covalently attached chondroitin sulfate glycosaminoglycan chains. NG2 also inhibited neurite growth from embryonic rat dorsal root ganglia neurons on substrates containing laminin. However, when the sensory neurons were plated onto surfaces containing the L1 glycoprotein and NG2, neurite growth was not inhibited. These results demonstrate that the NG2 proteoglycan provides an unfavorable substrate for axonal growth. Cells that express this proteoglycan in vivo may participate in axonal guidance by defining areas of the developing CNS that are nonpermissive for axonal extension from specific classes of developing neurons.
Nitric oxide synthase in Muller cells and neurons of salamander and fish retina
Authors: Liepe BA , Stone C , Koistinaho J , Copenhagen DR
Published: April 2018

Nitric oxide synthase (NOS) is the biosynthetic enzyme of the signaling molecule nitric oxide (NO). NO donors have been reported to modulate conductances in cell types throughout the retina, from photoreceptors to ganglion cells. Previously, NOS immunoreactivity has been reported in amacrine cells and cells within the ganglion cell layer. Here, we have examined the cellular localization of NOS in the retinas of salamander, goldfish, and catfish using both an affinity-purified antiserum to brain NOS and NADPH diaphorase (NADPHd) histochemistry. These markers indicate that an NOS-like enzyme is localized not only to presumptive amacrine cells but also, depending on the species, to photoreceptor ellipsoids, to somata within the ganglion cell layer, and to horizontal cells. In addition to these neurons, our results indicate that Muller cells, the radial glia of the retina, also contain an NOS- like enzyme. In support of this latter conclusion, cells morphologically similar to Muller cells were positive for NADPHd staining in all three species. In salamander, NOS-like immunoreactivity, NADPHd staining, and binding of anti-GFAP (a marker for glia) were localized to cells that were morphologically indistinguishable from Muller cells. In goldfish, reactivity to both anti-NOS and anti-vimentin (a marker for glia) colocalized to radial processes extending through the inner retina to the inner limiting membrane. These observations are the first to indicate the presence of an NOS-like enzyme in Muller cells and suggest that these glia could be a ready source of NO for target neurons throughout the retina.
The cortical component of experience-dependent synaptic plasticity in the rat barrel cortex
Authors: Fox K
Published: April 2018

Rats were raised with altered tactile experience from P0 by removing all but one vibrissa (D1) from one side of the face (D1-spared animals). This procedure (univibrissa rearing) has previously been shown to cause neurons in cortical barrels surrounding D1 to develop greater than normal responses to D1 vibrissa stimulation and smaller than normal responses to principal vibrissa stimulation (Fox, 1992). In this study, it was found that the potentiated D1 responses could be attenuated by acute microlesions placed in the D1 barrel, while principal vibrissa responses were unchanged or even slightly elevated for the same cases. The ratio of the average D1 to principal vibrissa response was approximately proportional to the volume of tissue damaged in the D1 barrel. This result implies that the synaptic plasticity seen in cortex of D1-spared animals is due to synaptic changes that take place within the barrel cortex rather than to relay of changes occurring at a subcortical level. In addition, lesions aimed at the septum between D1 and an adjacent barrel almost completely abolished responses to D1 stimulation in that barrel, including short-latency responses (5–10 msec). Only neurons severed horizontally from D1 were affected. Neurons that maintained a connection with the D1 barrel via a bridge of septal tissue preserved their usual elevated levels of response to D1 stimulation and their aberrant short-latency responses. This result implies that pathways radiating out from the D1 barrel/column, and connecting neurons in the D1 barrel to cells in surrounding barrels, undergo synaptic plasticity induced by univibrissa rearing.
Essential role of myosin light chain kinase in the mechanism for MgATP- dependent priming of exocytosis in adrenal chromaffin cells
Authors: Kumakura K , Sasaki K , Sakurai T , Ohara-Imaizumi M , Misonou H , Nakamura S , Matsuda Y , Nonomura Y
Published: April 2018

Ca(2+)-induced exocytosis in chromaffin cells now seems to consist of at least two distinct steps:MgATP-dependent Ca(2+)-dependent priming of the secretory apparatus, and Ca(2+)-dependent MgATP-independent step that triggers exocytosis (Bittner and Holz, 1992). Recently we found that a specific inhibitor of myosin light chain kinase (MLCK), wortmannin, inhibits Ca(2+)-induced catecholamine release from digitonin-permeabilized chromaffin cells, suggesting an implication of MLCK in the mechanisms of Ca(2+)-induced exocytosis (Imaizumi et al., 1992b). To elucidate further the implication of MLCK in the mechanism of exocytosis, we studied the effects of wortmannin and a peptide inhibitor (SM-1) corresponding to the pseudosubstrate domain of MLCK on MgATP-dependent and MgATP-independent release in digitonin- permeabilized chromaffin cells. Ca(2+)-induced exocytosis from the permeabilized cells in the presence of MgATP was inhibited by both SM-1 and wortmannin. Inhibitory effect of wortmannin on the rate of release induced by 10 microM Ca2+ in the presence of MgATP was much prominent in the later phase (1–10 min), although the initial rate was also decreased. SM-1 strongly inhibited ATP-dependent release without affecting Ca(2+)-dependent ATP-independent release at all. In addition, priming effect of MgATP that underlies Ca(2+)-dependent ATP-independent release was remarkably reduced by both wortmannin and SM-1. These results suggest that MLCK plays an essential role in ATP-dependent priming of Ca(2+)-induced exocytosis in chromaffin cells.
Propofol modulates activation and desensitization of GABAA receptors in cultured murine hippocampal neurons
Authors: Orser BA , Wang LY , Pennefather PS , MacDonald JF
Published: April 2018

Propofol (2,6 di-isopropylphenol) is an alkyphenol recently introduced for use as a general anesthetic. The modulation of GABAA receptor activation and desensitization by propofol was studied using a rapid perfusion system and whole-cell voltage-clamp recordings from mouse hippocampal neurons. The effects of concentrations of propofol used clinically on single-channel and synaptic currents were also examined. Propofol evoked current responses (EC50 = 61 microM) and shifted the dose-response curve of GABA-activated current to the left without altering the maximum of the GABA response. Preincubation with propofol and GABA led to desensitization of the GABA response (EC50 = 454 microM and 23 microM, respectively). Saturating concentrations of GABA (600 microM) evoked currents that peaked and then declined in a biexponential fashion with fast and slow time constants of tau f = 1.0 sec and tau s = 3.5 sec. Propofol (10 microM) did not change the amplitude of the peak response but decreased the rates of decay approximately 1.5-fold and enhanced the steady-state current proportionately. Recovery from desensitization was also biexponential (tau f = 11 sec, tau s = 69 sec) but not influenced by propofol. Single- channel recordings from outside-out patches demonstrated that both propofol and GABA activated channels with a 30 pS and 21 pS open state. Propofol increased the frequency but not the duration or conductance of GABA-activated events. Miniature inhibitory postsynaptic currents (mlPSCs) were evoked by the application of hypertonic sucrose to the cell soma. Propofol (2 microM) prolonged the decay time of mlPSCs to an extent similar to which it increased the open probability of GABA- activated channels (2.3- vs 3-fold). A sequential model, based on a previous scheme of GABA receptor gating (Weiss and Magelby, 1989), is presented to summarize propofol's actions on GABAA receptor function. We show through simulation that the model reliably reproduced the whole- cell tracings. Our results indicate that propofol's neurodepressive actions will be associated with enhancement of inhibitory synaptic transmission.