bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantitative study of tyrosine hydroxylase protein levels within the somatic area of the rat locus coeruleus during postnatal development
Authors: Bezin L , Marcel D , Rousset C , Pujol JF , Weissmann D
Published: April 2018

To date only global dosages of tyrosine hydroxylase (TH) protein have been realized in the locus coeruleus (LC) without discriminating the enzyme contained in the cell body area from the one in the surrounding neuropil. The preceding immunohistochemical study (Bezin et al., 1994) revealed a dramatic plasticity of the cellular expression of TH in the LC during the postnatal development of the rat. It was therefore necessary to develop a quantitative biochemical approach, strengthened by a great anatomical resolution, to follow the developmental evolution of TH levels exactly in the space containing the coerulean TH- immunoreactive perikarya. In the present work two biochemical parameters necessary for precisely defining the phenotypic characterization of TH expression within the rat LC have been established during the postnatal development at six different stages: postnatal day 4 (PND4), PND10, PND14, PND21, PND30, and PND42. TH tissue concentration and content were precisely determined along the caudorostral extent of the LC within the previously (Bezin et al., 1994) defined spaces delimited by the TH-containing perikarya. TH tissue concentration remained quite stable during the postnatal development. TH quantity exhibited few age-related variations with a transient peak at PND10 and followed the same evolution as the volume containing the TH-expressing perikarya. The mean cell contribution to the total quantity of TH measured in the whole LC showed important age- related fluctuations with a dramatic peak at PND10 followed by a drastic decrease until PND21.(ABSTRACT TRUNCATED AT 250 WORDS)
The computation of position sense from spindles in mono- and multiarticular muscles
Authors: Scott SH , Loeb GE
Published: April 2018

It is known that muscle spindles provide the majority of information about limb position, but little is known about how position sense is computed from their signals. We have developed a family of musculoskeletal models in order to determine some of the fundamental properties associated with transforming noisy spindle information into putative internal coordinate frames for position sense. A two-joint model was developed containing one biarticular and two monoarticular muscles with a total of 1000 sensors distributed among them. The sensors were assumed to function like spindle secondary afferents under fusimotor control designed to optimize their ability to encode static position in the presence of constant output noise. The optimal distribution of sensors was found to depend strongly on the coordinate frame in which position was measured (intersegmental angle, segment orientation, or end-point of the limb) and on the topology of the biarticular muscle with respect to the plane of motion. A similar analysis was performed for an anthropometric model of the human arm, using previously published counts of muscle spindles. In general, the actual distribution of spindles about the elbow and shoulder does not seem to favor any single coordinate frame for position sense. We also looked at the potential accuracy in detecting changes in joint angles based on the distribution of muscle spindles throughout the human body. The distribution of spindles about individual joints accounts well for psychophysical data showing a proximodistal descending gradient of angular resolution that partially reflects the relative importance of more proximal joints for determining the location of the end-point.
Neuronal and non-neuronal aromatase in primary cultures of developing zebra finch telencephalon
Authors: Schlinger BA , Amur-Umarjee S , Shen P , Campagnoni AT , Arnold AP
Published: April 2018

Estrogenic metabolites of circulating androgens have important effects on the organization and activation of neural circuits controlling reproductive behavior and physiology in males of many vertebrate species. Previous studies indicate that aromatase, the enzyme that converts androgens to estrogens, is expressed most abundantly in neurons in limbic brain regions. Songbirds are unique in that aromatase is expressed at unusually high levels throughout the telencephalon of both males and females. We assume that estrogens formed in the telencephalon itself masculinize neural circuits controlling song, since the brain is a major source of circulating estrogens in adult males. However, the cellular localization of telencephalic aromatase in songbirds remains unknown. We have established primary cultures from telencephalons of developing zebra finches and found aromatase activity (conversion of 3H-androstenedione or 3H-testosterone to 3H-estrone plus 3H-estradiol) at some of the highest levels reported for brain tissue of any species. Both neurons and glia were identified in these cultures based on cell morphology and labeling by specific immunohistochemical markers. However, when culture conditions were manipulated to reduce the incidence of either neurons or glia by varying the age of cultures or their plating density, treating with the neurotoxin kainic acid, physically shaking off loosely attached neurons, or preparing cultures in media that encouraged enrichment of neurons, high levels of aromatase persisted. Furthermore, Northern blot analysis of total RNA extracted from enriched neuronal or glial cultures indicated the presence of aromatase mRNA in both cell preparations. In situ hybridization with a zebra finch aromatase cDNA probe conjugated to digoxigenin showed the cultures contained darkly labeled neurons and lightly labeled non-neuronal cells, presumably astrocytes. We conclude that aromatase is expressed in both neuronal and non-neuronal cells in these cultures, suggesting that both cell types may also express the enzyme in vivo. The presence of aromatase outside of neurons suggests that glia may be targets of estrogen action or that glia may supply some estrogen to the estrogen-sensitive neural circuits in this species.
Opioid receptors modulate diverse types of calcium channels in the nucleus tractus solitarius of the rat
Authors: Rhim H , Miller RJ
Published: April 2018

We have investigated the coupling between opioid receptors and different types of Ca2+ channels in neurons acutely isolated from the nucleus tractus solitarius (NTS) of the rat. Using fura-2-based imaging we found that Ca2+ transients evoked by depolarization with 50 mM KCl were suppressed by the mu-opioid receptor agonist D-Ala2,N-MePhe4,Gly5- ol-enkephalin (DAMGO) and less effectively by the kappa-receptor agonist U-69,593. The delta-receptor agonist D-Pen2,D-Pen5-enkephalin (DPDPE) was ineffective. In whole-cell voltage-clamp recordings from these neurons, depolarizing voltage steps elicited high-threshold Ca2+ currents that could be distinguished pharmacologically into different components. Part of the current could be blocked by dihydropyridines, part by omega-conotoxin-GVIA and part by omega-agatoxin-IVA. This suggests that the neurons contained L-, N-, and P/Q-type Ca2+ channels. DAMGO and U-69,593 both blocked part of the Ca2+ current but DPDPE was ineffective. Perfusion of GTP-gamma-S into the cells produced a rapid rundown of the Ca2+ current and occluded further effects of the opioid agonists, suggesting the involvement of a G-protein in the coupling mechanism. Inhibition of L-channels did not alter the effect of DAMGO. On the other hand inhibition of N-channels occluded about 80% of the effect of DAMGO. Inhibition of the P/Q-current occluded the remainder of the DAMGO effect. Thus, it appears that activation of opioid receptors can inhibit N- and P/Q-type Ca2+ channels but not L-channels in these cells. It is likely that such effects are important in opioid- mediated inhibition of transmitter release in the brain.
Estradiol regulates hippocampal dendritic spine density via an N-methyl- D-aspartate receptor-dependent mechanism
Authors: Woolley CS , McEwen BS
Published: April 2018

In the adult female rat, the densities of dendritic spines and synapses on hippocampal CA1 pyramidal cells are dependent upon the ovarian steroid estradiol; moreover, spine and synapse density fluctuate naturally as ovarian steroid levels vary across the estrous cycle. To determine whether the effects of estradiol on dendritic spine density require activation of specific neurotransmitter systems, we have treated animals concurrently with estradiol and one of four selective neurotransmitter receptor antagonists: MK 801, a noncompetitive NMDA receptor antagonist; CGP 43487, a competitive NMDA receptor antagonist; NBQX, an AMPA receptor antagonist; or scopolamine, a muscarinic receptor antagonist. Our results indicate that the effects of estradiol can be blocked by treatment with either of the NMDA receptor antagonists, but treatment with an AMPA or muscarinic receptor antagonist has no effect on spine density. Thus, we have concluded that estradiol exerts its effect on hippocampal dendritic spine density via a mechanism requiring activation specifically of NMDA receptors.
Facilitation of calcium-dependent potassium current
Authors: Thompson SH
Published: April 2018

The activation of Ca-dependent K+ current, Ic, was studied in macropatches on the cell bodies of molluscan neurons. When a depolarizing voltage-clamp pulse was applied repeatedly, Ic facilitated in a manner that resembled the facilitation of synaptic transmitter release. Facilitation was characterized by an increase in Ic amplitude, a progressive increase in instantaneous outward current, and a decrease in utilization time. Experiments were done to investigate the mechanism responsible for Ic facilitation. Facilitation was reduced by microinjection of an exogenous Ca2+ buffer into the cytoplasm, indicating that facilitation is a Ca(2+)-dependent process. It was also reduced at elevated temperatures. Conversely, facilitation was greatly potentiated by blocking the Na/Ca exchange mechanism. It is concluded that the facilitation of Ca-dependent K+ current results from the accumulation of Ca2+ at the inner face of the membrane during the repeated activation of Ca2+ channels by depolarization. The Ca2+ indicator fluo-3 was used in fluorescence imaging experiments to measure changes in [Ca]i near the cell membrane during repeated depolarizing pulses and the interpretation of these results was aided by numerical simulations of Ca2+ accumulation, diffusion, and buffering in the peripheral cytoplasm. These experiments showed that the time course of Ic facilitation matches the time course of Ca2+ accumulation at the membrane. It was found that the strength of Ic facilitation varies among patches on the same neuron, suggesting that the accumulation of Ca2+ is not uniform along the inner surface of the membrane and that gradients in [Ca]i develop and are maintained during trains of depolarizing pulses. Potential mechanisms that may lead to local differences in Ca2+ accumulation and Ic facilitation are discussed.
Sharp wave-associated high-frequency oscillation (200 Hz) in the intact hippocampus: network and intracellular mechanisms
Authors: Ylinen A , Bragin A , Nadasdy Z , Jando G , Szabo I , Sik A , Buzsaki G
Published: April 2018

Sharp wave bursts, induced by a cooperative discharge of CA3 pyramidal cells, are the most synchronous physiological pattern in the hippocampus. In conjunction with sharp wave bursts, CA1 pyramidal cells display a high-frequency (200 Hz) network oscillation (ripple). In the present study extracellular field and unit activity was recorded simultaneously from 16 closely spaces sites in the awake rat and the intracellular activity of CA1 pyramidal cells during the network oscillation was studied under anesthesia. Current source density analysis of the high-frequency oscillation revealed circumscribed sinks and sources in the vicinity of the pyramidal layer. Single pyramidal cells discharged at a low frequency but were phase locked to the negative peak of the locally derived field oscillation. Approximately 10% of the simultaneously recorded pyramidal cells fired during a given oscillatory event. Putative interneurons increased their discharge rates during the field ripples severalfold and often maintained a 200 Hz frequency during the oscillatory event. Under urethane and ketamine anesthesia the frequency of ripples was slower (100–120 Hz) than in the awake rat (180–200 Hz). Halothane anesthesia prevented the occurrence of high-frequency field oscillations in the CA1 region. Both the amplitude (1–4 mV) and phase of the intracellular ripple, but not its frequency, were voltage dependent. The amplitude of intracellular ripple was smallest between -70 and -80 mV. The phase of intracellular oscillation relative to the extracellular ripple reversed when the membrane was hyperpolarized more than -80 mV. A histologically verified CA1 basket cell increased its firing rate during the network oscillation and discharged at the frequency of the extracellular ripple. These findings indicate that the intracellularly recorded fast oscillatory rhythm is not solely dependent on membrane currents intrinsic to the CA1 pyramidal cells but it is a network driven phenomenon dependent upon the participation of inhibitory interneurons. We hypothesize that fast field oscillation (200 Hz) in the CA1 region reflects summed IPSPs in pyramidal cells as a result of high-frequency barrage of interneurons. The sharp wave associated synchronous discharge of pyramidal cells in the millisecond range can exert a powerful influence on retrohippocampal targets and may facilitate the transfer of transiently stored memory traces from the hippocampus to the entorhinal cortex.
Gamma (40-100 Hz) oscillation in the hippocampus of the behaving rat
Authors: Bragin A , Jando G , Nadasdy Z , Hetke J , Wise K , Buzsaki G
Published: April 2018

The cellular generation and spatial distribution of gamma frequency (40– 100 Hz) activity was examined in the hippocampus of the awake rat. Field potentials and unit activity were recorded by multiple site silicon probes (5- and 16-site shanks) and wire electrode arrays. Gamma waves were highly coherent along the long axis of the dentate hilus, but average coherence decreased rapidly in the CA3 and CA1 directions. Analysis of short epochs revealed large fluctuations in coherence values between the dentate and CA1 gamma waves. Current source density analysis revealed large sinks and sources in the dentate gyrus with spatial distribution similar to the dipoles evoked by stimulation of the perforant path. The frequency changes of gamma and theta waves positively correlated (40–100 Hz and 5–10 Hz, respectively). Putative interneurons in the dentate gyrus discharged at gamma frequency and were phase-locked to the ascending part of the gamma waves recorded from the hilus. Following bilateral lesion of the entorhinal cortex the power and frequency of hilar gamma activity significantly decreased or disappeared. Instead, a large amplitude but slower gamma pattern (25–50 Hz) emerged in the CA3-CA1 network. We suggest that gamma oscillation emerges from an interaction between intrinsic oscillatory properties of interneurons and the network properties of the dentate gyrus. We also hypothesize that under physiological conditions the hilar gamma oscillation may be entrained by the entorhinal rhythm and that gamma oscillation in the CA3-CA1 circuitry is suppressed by either the hilar region or the entorhinal cortex.
Enduring effects of chronic corticosterone treatment on spatial learning, synaptic plasticity, and hippocampal neuropathology in young and mid-aged rats
Authors: Bodnoff SR , Humphreys AG , Lehman JC , Diamond DM , Rose GM , Meaney MJ
Published: April 2018

Prolonged treatment with stress levels of corticosterone has been reported to produce changes in the hippocampus. In the experiments reported here, we examined for functional and morphological consequences of this treatment. First, young adult or mid-aged male Long-Evans rats were treated for either 1 or 3 months with corticosterone, at a dose sufficient to mimic the elevated hormone levels observed following exposure to mild stress. Two weeks following the termination of treatment, the animals were tested in the Morris water maze to assess spatial learning. No behavioral deficits were observed after 1 month of treatment. A 3 month treatment period also had no effect in young rats, but produced a learning impairment in the mid-aged rats. We then examined whether the effect of elevated corticosterone in mid-aged animals could be produced by a physiological stressor. Mid-aged rats were maintained for 6 months under conditions of low or high social stress. Six months of exposure to high social stress produced significant spatial learning impairments in the Morris water maze. These effects were absent in high social stress animals that had been previously adrenalectomized (with low-level corticosterone replacement), suggesting that elevated glucocorticoid levels mediate the effects of stress on spatial memory in older animals. In a final experiment, mid-aged rats were treated with corticosterone at levels that mimicked those naturally occurring at the diurnal peak (medium-B: 12–17 micrograms/dl) or in response to stress (high-B: 25–32 micrograms/dl). Only rats exposed to high levels of corticosterone demonstrated impaired performance in the Morris water maze.(ABSTRACT TRUNCATED AT 250 WORDS)
Head direction cells recorded in the anterior thalamic nuclei of freely moving rats
Authors: Taube JS
Published: April 2018

Metabotropic glutamate receptors (mGluRs) are critically involved in the maintenance of long-term potentiation (LTP) (Reymann and Matthies, 1989; Behnisch et al., 1991; Izumi et al., 1991; Bashir et al., 1993). In order to assess further the physiological role of MGluRs in LTP, we injected freely moving rats with the recently available, competitive mGluR antagonist (R,S)-alpha-methyl-4-carboxyphenylglycine (MCPG) intraventricularly and recorded extracellularly the population spike (PS) as well as the field excitatory postsynaptic potential (fEPSP) of the granule cells of the dentate gyrus in response to stimulation of fibers of the perforant path. MCPG was administered in two concentrations (A = 20 mM/5 microliters; B = 200 mM/5 microliters) either 30 min prior to or 5 min after LTP induction. Sodium chloride infusion served as a control. Normal synaptic transmission was not altered by MCPG. However, the mGluR antagonist inhibited LTP in a concentration-dependent manner. Concentration A did not influence the potentiation shortly after the tetanus. In the PS, short-term potentiation (STP), which is decremental in its time course, occurred normally, but in contrast to controls the potentiation declined back to baseline values after 2–3 hr. This dose also reduced the posttetanic increase in the slope function of the fEPSP, and led to a time course of potentiation similar to that for the PS. Concentration B completely abolished the tetanus-induced potentiation. This block was similar to that obtained for the NMDA antagonist 2-amino-5-phosphonopentanoate (AP5). Both MCPG concentrations had no influence on the time course of preestablished LTP. These effects seem to be due to the action of the (+)-isomer of MCPG, since intracerebroventricular application of the (- )-isomer was without effect on the duration and magnitude of LTP. In addition, we were interested in the mGluR subtypes involved in the blocking mechanism of MCPG. 1S,3R-aminocyclopentane-1,3-dicarboxic acid (ACPD)-activated PPI hydrolysis in hippocampal slices was competitively inhibited by MCPG at a concentration of 1 mM or higher. In contrast, this concentration of MCPG did not affect the reduction of forskolin- stimulated cAMP formation by ACPD. These results corroborate recent findings that mGluRs are required for the induction of LTP in CA1 and CA3 in vitro (Bashir et al., 1993; Sergueeva et al., 1993) and in vivo (Riedel and Reymann, 1993). The process of STP is found to be independent of mGluR activation.(ABSTRACT TRUNCATED AT 400 WORDS)
Cytotoxic effects of sigma ligands: sigma receptor-mediated alterations in cellular morphology and viability
Authors: Vilner BJ , de Costa BR , Bowen WD
Published: April 2018

The morphological effects of several neuroleptics as well as other novel and prototypic sigma ligands were examined by addition to cultures of C6 glioma cells. Sigma ligands caused loss of processes, assumption of spherical shape, and cessation of cell division. The time course and magnitude of this effect were dependent on the concentration of sigma ligand. Continued exposure to sigma compounds ultimately resulted in cell death. However, the morphological effect was reversible when sigma ligand was removed shortly after rounding. The potency of compounds to produce these effects generally correlated with binding affinity at sigma receptors of C6 glioma cell membranes labeled with [3H](+)-pentazocine. At a concentration of 100 microM, haloperidol, reduced haloperidol, fluphenazine, perphenazine, trifluoperazine, BD737, LR172, BD1008, and SH344 produced significant effects in 3–6 hr of exposure. Other compounds, such as trifluperidol, thioridazine, and (-)-butaclamol, produced significant effects by 24 hr of exposure. Despite the requirement of micromolar concentrations of ligand (some compounds were effective at 30 microM), the effect showed a remarkable specificity for compounds exhibiting sigma receptor binding affinity. Neuroleptics lacking potent sigma affinity [e.g., (-)- sulpiride, (+)-butaclamol, and clozapine] and other compounds that lack significant sigma affinity but that are agonists or antagonists at dopamine, serotonin, adrenergic, glutamate, phencyclidine, GABA, opiate, or muscarinic cholinergic receptors were without effect on cellular morphology at concentrations up to 300 microM over a period of 72 hr. Likewise, blockers and activators of Na+, K+, and Ca2+ channels and a monoamine oxidase inhibitor devoid of sigma affinity were without effect. Interestingly, 1,3-di-o-tolylguanidine (DTG), (+)-3-(3- hydroxyphenyl)-N-(1-propyl)piperidine [(+)-3-PPP], (+)-pentazocine, (+)- cyclazocine, and other sigma-active benzomorphans and morphinans appeared inactive in up to 72 hr of culture. However, these compounds interacted synergistically with a subeffective dose of BD737 (30 microM) to produce effects usually in 6 hr or less. Also, the pH of the culture medium had a profound effect on the activity of sigma compounds. Increasing the pH from the normal range of 7.2–7.4 to pH 8.3– 8.5 shifted the dose curves (30, 100, 300 microM) for all sigma compounds to the left. Under these conditions, DTG, (+)-3-PPP, and benzomorphans produced effects in 24 hr or less.(ABSTRACT TRUNCATED AT 400 WORDS)
Activation of a metabotropic glutamate receptor increases intracellular calcium concentrations in neurons of the avian cochlear nucleus
Authors: Zirpel L , Lachica EA , Rubel EW
Published: April 2018

Metabotropic glutamate receptors have been shown to stimulate phosphatidylinositol metabolism, and subsequently liberate Ca2+ from intracellular stores, in a variety of tissue and cell types. We previously demonstrated that glutamate could stimulate phosphatidylinositol metabolism, generating inositol-1,4,5- trisphosphate (IP3), in isolated cochlear nucleus tissue from the chick. Using the calcium indicator dye fura-2 and ratiometric fluorescent imaging, this study examined the ability of glutamate and its analogs to liberate Ca2+ from intracellular stores of neurons of the avian cochlear nucleus, and qualitatively characterized the pharmacological profile of such an action. In normal, Ca(2+)-containing medium, glutamate, kainate (KA), alpha-amino-3-hydroxy-5-methyl- isoxazole-4-propionate (AMPA), NMDA, quisqualate (QUIS), and (+/-)- aminocyclopentane-trans-dicarboxylate (ACPD) elicited increases in intracellular calcium concentrations ([Ca2+]i). In the absence of external Ca2+, glutamate, quisqualate, and ACPD evoked increases in [Ca2+]i. In normal medium, the ionotropic glutamate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and the NMDA receptor antagonist 2-amino-5-phosphonovalerate (APV) attenuated but did not abolish the glutamate-evoked response and had no effect on the ACPD- evoked response. The putative metabotropic glutamate receptor antagonist 2-amino-3-phosphonopropionate (AP3) was without effect on the glutamate- and ACPD-evoked increases in [Ca2+]i in Ca(2+)-free medium. We conclude that a metabotropic glutamate receptor (mGluR) is present on cochlear nucleus neurons and is able to stimulate the phosphatidylinositol metabolism--Ca2+ signal transduction cascade.
Expression and antibody inhibition of P-type calcium channels in human small-cell lung carcinoma cells
Authors: Barry EL , Viglione MP , Kim YI , Froehner SC
Published: April 2018

P-type channels, a recently described form of voltage-gated calcium channels, are found in many central and peripheral neurons. In the present study, a partial cDNA clone sharing extensive nucleotide identity with a putative P-type voltage-gated calcium channel alpha 1 subunit was isolated from a small-cell lung carcinoma (SCLC) cell line. Anti-peptide antibodies generated to a unique acidic stretch in the IVS5-S6 linker region of the putative SCLC P-type channel reacted specifically with a SCLC fusion protein produced in bacteria and with a cell surface molecule in SCLC cells. Calcium currents in SCLC cells, measured by whole-cell patch clamp, were inhibited by these antibodies and by the P-type channel-specific toxin omega-agatoxin IVA. The inhibitory effects of the antibody and the toxin were not additive, consistent with their proposed action on the same type of channel. These results provide evidence for the expression of P-type calcium channels by SCLC cells. The expression of neuron-related molecules by these cells is of particular interest because small-cell lung carcinoma is frequently associated with paraneoplastic disorders affecting the nervous system.
Muscular production of individuated finger movements: the roles of extrinsic finger muscles
Authors: Schieber MH
Published: April 2018

Individuated finger movements--those in which one or more fingers are moved relatively independently of the movement or posture of other body parts--are produced in part by the action of the extrinsic finger muscles. Flexion/extension movements of the fingers are particularly dependent on these extrinsic muscles, most of which are multitendoned. How can contraction of multitendoned muscles move one digit without producing equivalent motion in other digits? This question was addressed by recording EMG activity from muscles of the forearm as trained rhesus monkeys performed flexion and extension individuated movements of each digit of the hand and of the wrist. Recordings showed that during movements of different fingers, a given muscle could act as an agonist, antagonist, or stabilizer of the digits it serves. Furthermore, during a given finger movement, several different muscles typically were active. A three-level connection model was developed that computed the relative motion of the digits during each finger movement based on the changes in EMG activity in the recorded muscles. The model showed that EMG activity changes in the extrinsic finger muscles, and the thenar muscles, could account for most of the motion of both the instructed digit and noninstructed digits. These results indicate that individuated finger movements were produced not by independent sets of muscles acting on each digit, but by the activity of several muscles, many of which act on more than one digit, combined such that the net effect was movement of one digit more than others.
Novel classes of responsive and unresponsive C nociceptors in human skin
Authors: Schmidt R , Schmelz M , Forster C , Ringkamp M , Torebjork E , Handwerker H
Published: April 2018

One hundred ninety-four cutaneous C-fibers were recorded from the human peroneal nerve; 118 units were found by mechanical stimuli and 76 units were detected by electrical stimulation through a surface electrode. Needle electrodes were then inserted for electrical intradermal stimulation in the innervation territory of the units. Afferent and efferent sympathetic C-fibers were identified by slowing of conduction velocity after activation either by physical or chemical skin stimuli, or by arousal maneuvers eliciting sympathetic reflexes. In addition to mechano-heat-responsive C units (CMH) also found in previous studies, we here report on novel classes of C nociceptors in human skin, namely, units responding only to mechanical stimuli (CM), units responding only to heating (CH), and units that were insensitive to mechanical and heating stimuli and also to sympathetic provocation tests (CMiHi). With the electrical search technique we found 45% CMH, 13% CM, 6% CH, 24% CMiHi, and 12% sympathetic units. Excitation by topically applied mustard oil occurred in 58% of CMH units, and in one-third of CM and CMiHi units, respectively. Some CM, CH, and CMiHi units were sensitized to heating and/or to mechanical stimuli after topical application of mustard oil or capsaicin. These units then acquired responsiveness to a stimulus modality to which they previously were insensitive. Such recruitment of previously silent nociceptors implies spatial summation to the nociceptive barrage at central levels, and may contribute both to primary hyperalgesia to heat and pressure after chemical irritation, and to secondary hyperalgesia as a consequence of central sensitization.
Monopolar cell axons in the first optic neuropil of the housefly, Musca domestica L., undergo daily fluctuations in diameter that have a circadian basis
Authors: Pyza E , Meinertzhagen IA
Published: April 2018

Two types of monopolar cell interneurons, each with a single representative in every unit cartridge of the first optic neuropil, or lamina, of the housefly's optic lobe, have axons that undergo cyclical changes in diameter. The axons are largest during the beginning of day in a normal LD light cycle and smallest during the middle of the night, changes that were however significant only for one of the cells (L2). The axon cross-sectional area and its cyclical change for both L1 and L2 were both larger in the proximal lamina. The changes are not a simple consequence of relative osmotic change. Dehydration paradoxically increases axon size, and also fails to alter the day/night rhythm of axon size changes. Under conditions of constant darkness, both axons decrease in size, and one of the cells (L2) retains its cyclical size changes, being larger in the subjective day than in the subjective night. Under conditions of constant light, both axons increase in size, and L2 again shows a cyclical size change, just as under conditions of constant darkness. These changes seen under constant conditions are, by definition, circadian in origin. The effects of exposure to light or darkness can partially reset these circadian changes. One extra hour of light during the day increases the size of L1 and L2, whereas 1 hr of extra dark during the night does not decrease their size. It takes 13 hr of light to reverse the rhythm in size. The mechanism for all these changes is unclear but may involve ionic fluxes, possibly that are secondary to osmotic shifts and probably that involve at least two independent processes.
Dynamics of protein phosphorylation in sensory neurons of Aplysia
Authors: Homayouni R , Byrne JH , Eskin A
Published: April 2018

Protein phosphorylation plays important roles in the mechanisms underlying serotonin (5-HT)-induced presynaptic facilitation of Aplysia sensory neurons. To study mechanisms involved in facilitation, we investigated the pattern of protein phosphorylation in sensory neurons as a function of different durations of 5-HT. Two minutes and 1.5 hr treatments with 5-HT altered the phosphorylation of 5 and 10 proteins, respectively. These different duration treatments with 5-HT produced unique effects on the phosphorylation of different sets of proteins. This result suggests that cells may encode and measure the duration of a stimulus by the pattern of specific proteins that are phosphorylated or dephosphorylated. In addition, because the changes in phosphorylation produced by 2 min treatments with 5-HT were not observed after 25 min treatments with 5-HT, mechanisms must exist for the transient phosphorylation of some proteins even when the 5-HT treatment persists. Anisomycin, an inhibitor of protein synthesis, blocked the effect of 1.5 hr treatments with 5-HT on the phosphorylation of six proteins but had no effect on the phosphorylation change of four other proteins. Both CPT-cAMP (an activator of protein kinase A) and PDAc (an activator of protein kinase C) mimicked the effects of 5-HT on four proteins. Interestingly, the effect of 5-HT on these four proteins did not require protein synthesis. CPT-cAMP, but not PDAc, mimicked the effect of 5-HT on one protein (L55) and, the effect of 5-HT on this protein appeared to require protein synthesis. Because both activation of PKA and protein synthesis are involved in the induction of long-term facilitation, protein L55 is a good candidate for a protein that might play a key role in long-term facilitation. Finally, the effects of 5-HT on four proteins were not mimicked by either CPT-cAMP or PDAc. This finding raises the interesting possibility that some effects of 5-HT are mediated by second-messenger systems other than PKA or PKC.
Retrograde transport of plasticity signals in Aplysia sensory neurons following axonal injury
Authors: Gunstream JD , Castro GA , Walters ET
Published: April 2018

Following injury to their peripheral branches, mechanosensory neurons in Aplysia display long-term plasticity that is expressed as soma hyperexcitability, synaptic facilitation, and neurite outgrowth. To investigate the nature of signals that convey information about distant axonal injury, we have investigated the development of injury-induced soma hyperexcitability in two in vitro preparations. In isolated ganglia, proximal nerve crush caused hyperexcitability to appear sooner than did distal crush, and the difference in development of hyperexcitability indicated that the injury signal moved at a rate (36 mm/d) similar to previously reported rates of retrograde axonal transport in this animal. This hyperexcitability was not due to interruption of continuous retrograde transport of trophic substances (a negative signal) because inhibitors of axonal transport applied to uncrushed nerve segments did not induce hyperexcitability. Indeed, inhibitors of axonal transport blocked crush-induced hyperexcitability, indicating that positive injury signals are involved. Crush-induced hyperexcitability was unaffected by bathing the nerve in tetrodotoxin or the ganglion in Cd2+, suggesting that the retrograde signals depend upon neither spike activity in the nerve nor synaptic transmission in the ganglion. Close excision of sensory neuron somata (which largely eliminated delays attributable to axonal transport) produced soma hyperexcitability that was expressed after 10 hr and lasted at least 17 d. These data indicate that axonal injury mobilizes signal molecules that are conveyed by retrograde axonal transport into the soma and possibly the nucleus, where they induce long-term plasticity similar to that expressed by these cells during learning and memory.
Inhibition of voltage-gated K+ channel gene expression by the neuropeptide thyrotropin-releasing hormone
Authors: Takimoto K , Gealy R , Fomina AF , Trimmer JS , Levitan ES
Published: April 2018

Many neurotransmitters regulate action potential activity in neuronal, endocrine, and cardiac cells by rapidly modulating the gating of K+ channels. Neurotransmitters might also produce prolonged effects on excitability by regulating the expression of K+ channel genes. Here we show that the neuropeptide thyrotropin-releasing hormone (TRH) down- regulates Kv1.5 and Kv2.1 K+ channel mRNAs in clonal pituitary cells. The effect on Kv1.5 mRNA expression does not require protein synthesis and is due to decreased transcription. Immunoblots demonstrate that Kv1.5 and Kv2.1 immunoreactivities are significantly reduced by TRH within 12 hr. The change in channel protein expression is associated with a decrease in voltage-gated K+ currents. Thus, TRH enhances excitability by inhibiting K+ channel gene expression. Neuropeptide regulation of K+ channel gene expression may produce long-term changes in neuronal action potential activity and synaptic transmission.
Neuronal and glial prostaglandin D synthase isozymes in chick dorsal root ganglia: a light and electron microscopic immunocytochemical study
Authors: Vesin MF , Urade Y , Hayaishi O , Droz B
Published: April 2018

Homogenates of chick dorsal root ganglia (DRG) and in vitro cultures of DRG neurons are known to synthesize prostaglandin (PG) D2. To specify the PGD synthase isozymes controlling PGD2 synthesis in DRG and to identify the DRG cells responsible for this synthesis, we applied polyclonal antibodies raised against rat brain or rat spleen PGD synthase isozymes to vibratome or cryostat slices of DRG previously fixed with a formaldehyde-lysine-periodate mixture and permeabilized with Triton X-100. The immunoreactivity indicating rat spleen PGD synthase, a glutathione (GSH)-requiring enzyme, was located in satellite cells encompassing particular large neurons of class A and in Schwann cells myelinating and enwrapping their initial axonal segments. In contrast, the immunoreactivity of rat brain PGD synthase, a GSH- independent enzyme, was restricted to particular ganglion cell perikarya: 33% of the DRG neurons were immunostained for rat brain PGD synthase, including 2% of large class A neurons and 40% of small class B neurons. Only 3.3% of rat brain PGD synthase-immunoreactive small B neurons coexpressed substance P, indicating that the immunoreactive neurons belong to the B1 subclass. By electron microscopy, 71 of 72 immunoreactive DRG cells were identified as small B neurons of the B1 subclass, and 71 of 77 B1 neurons were immunoreactive for rat brain PGD synthase. These results demonstrate that PGD2 formation in DRG is regulated by two isozymes: the GSH-requiring isozyme located in satellite and Schwann cells and the GSH-independent isozyme-confined to small B1 neurons.
Clustering of Na+ channels and node of Ranvier formation in remyelinating axons
Authors: Dugandzija-Novakovic S , Koszowski AG , Levinson SR , Shrager P
Published: April 2018

Polyclonal antibodies were raised against a well conserved region of the vertebrate Na+ channel and were affinity purified for use in immunocytochemistry. Focal demyelination of rat sciatic axons was initiated by an intraneural injection of lysolecithin and Na+ channel clustering was followed at several stages of myelin removal and repair. At 1 week post-injection axons contained long, fully demyelinated regions. Na+ channel clusters appeared only at heminodes forming the borders of these zones, and at widely spaced isolated sites that may represent former nodes of Ranvier. Over the next few days proliferating Schwann cells adhered to axons and began to extend processes. Clusters of Na+ channels appeared at the edges of these structures. As the Schwann cells elongated, the clusters seemed to move with them, since they remained at edges and the distance between aggregates increased. Clusters associated with different Schwann cells ultimately approached each other and appeared to fuse. Na+ channels then coalesced further at these sites, forming new nodes of Ranvier in regions that previously were internodal. If Schwann cell proliferation were blocked by mitomycin, no new clusters of Na+ channels appeared within internodes. Under these conditions, heminodal clusters remained visible at 1 week postinjection, but by 2 weeks they were no longer detectable, suggesting that proliferating Schwann cells are required for their maintenance. Clusters at normal nodes of Ranvier remained. It is concluded that Na+ channel aggregation and mobility in demyelinated nerve fibers is controlled by adhering Schwann cells, resulting in the formation of stable new nodes of Ranvier during remyelination.
An anti-CD9 monoclonal antibody promotes adhesion and induces proliferation of Schwann cells in vitro
Authors: Hadjiargyrou M , Patterson PH
Published: April 2018

We have recently found that CD9, a cell surface glycoprotein involved in intercellular signaling in hematopoietic cells, is also expressed by neurons and glia in the peripheral nervous system. Antibody perturbation experiments were conducted to examine the function of CD9 in neural cells. Three anti-CD9 monoclonal antibodies (mAbs) (ROCA1, ROCA2, B2C11) were tested for their ability to promote adhesion of several Schwann cell lines (S-16, RN22, JS1), primary Schwann cells and PC12 cells. Only B2C11 promotes adhesion in all cells tested. Although ROCA2 immunolabels living cells strongly, it had no effect on the adhesion of any of these cells. In addition, ROCA1 and several positive- staining, control mAbs also had no effect. Another mAb, 192-IgG, directed against the low affinity NGF receptor, also promotes the adhesion of S-16, PC12, and primary Schwann cells. In addition to adhesion, contact of S-16 Schwann cells with B2C11 specifically induces morphological changes and robust proliferation. None of the other mAbs, including 192-IgG, induce proliferation of S-16 cells. These results provide evidence that CD9 may be involved in signaling, activation and growth regulation of cells in the nervous system.
Chronic depolarization prevents programmed death of sympathetic neurons in vitro but does not support growth: requirement for Ca2+ influx but not Trk activation
Authors: Franklin JL , Sanz-Rodriguez C , Juhasz A , Deckwerth TL , Johnson EM
Published: April 2018

Continuous exposure of many types of neurons in cell culture to elevated concentrations of K+ greatly enhances their survival. This effect has been reported to be mediated by a sustained rise of cytoplasmic free Ca2+ concentration caused by influx of Ca2+ through voltage-gated channels activated by K(+)-induced chronic depolarization. In this report we investigate the effects of elevated K+ on the programmed death that embryonic rat sympathetic neurons undergo in culture when deprived of NGF. Elevated K+ in the culture medium did not significantly prevent death of NGF-deprived cells until after the third day following plating of embryonic day 21 neurons. On the fifth day after plating, incrementally increasing K+ concentrations in the culture medium from 5 to 100 mM caused chronic depolarization of neurons and had a biphasic effect on survival of NGF-deprived cells. Enhanced survival was steeply related to membrane potential, increasing from no enhanced survival in cells held at potentials between -51 and - 34 mV to 90–100% of control survival at about -21 mV. At potentials positive to -21 mV, survival decreased. Associated with the chronic depolarization was a sustained rise of steady-state free Ca2+ concentration that showed a biphasic relationship to membrane potential roughly similar to that exhibited by survival. Steady-state Ca2+ concentration increased with increasingly lower membrane potentials to a peak at about -23 mV (to approximately 240 nM from approximately 40 nM at about -51 mV) and then decreased at more positive potentials. The elevation of intracellular Ca2+ was largely blocked by dihydropyridine and phenylalkylamine Ca2+ channel antagonists and was potentiated by a dihydropyridine Ca2+ channel agonist. Neither the rise of Ca2+, or survival was affected by the Ca2+ channel antagonist, omega-conotoxin. Therefore, the Ca2+ elevation was probably caused by Ca2+ influx through L-type, but not N-type, channels. Antagonists of L channels blocked both survival and the sustained increase of steady-state free Ca2+ at similar concentrations, suggesting that the relevant factor determining survival of depolarized cells was Ca2+ influx rather than some other effect of depolarization. Surprisingly, however, there was no clear correlation between the sustained rise of Ca2+ and survival. Some membrane potentials that induced similar increases of Ca2+ concentration produced widely different levels of survival. While chronic depolarization promoted survival of neurons in the absence of NGF, cells supported in this manner showed little growth as measured by neurite extension, total cellular protein, and mean somal diameter.(ABSTRACT TRUNCATED AT 400 WORDS)
Immortalized murine striatal neuronal cell lines expressing dopamine receptors and cholinergic properties
Authors: Wainwright MS , Perry BD , Won LA , O'Malley KL , Wang WY , Ehrlich ME , Heller A
Published: April 2018

Immortalized hybrid cells were generated by somatic cell fusion of 18-d- old embryonic corpus striatum of the mouse strain C57BL/6J with the N18TG2 neuroblastoma. One of the cell populations obtained was treated with a combination of 1 mM n-butyric acid and 10 microM SKF 38393 (a specific D1 agonist), and a surviving cell population (E1X) was subcloned. Twenty-seven monoclonal cell lines were obtained and screened for the expression of striatal-specific characteristics including gamma-aminobutyric acid (GABA), choline acetyltransferase (ChAT), acetylcholine (ACh), mRNA for specific dopamine receptors, and dopamine- and adenosine 3′:5′-monophosphate-regulated phosphoprotein, M(r) 32,000 (DARPP-32), and functional D1 and D2 dopamine receptors. Neither the parent hybrid cell population (E1X) nor any of the monoclonal cell lines examined expressed GABA levels significantly different than that of the N18TG2 parent neuroblastoma cells (1.36 +/- 0.07 micrograms/mg protein). The range of ChAT activity in the monoclonal hybrid cell lines was 5.5 +/- 0.3 to 921.3 +/- 97.4 pmol/min/mg protein. Two of the cell lines expressing ChAT activity (X52 and X58) contained ACh (49.64 +/- 4.23 and 1.78 +/- 0.07 ng/mg protein, respectively). The neuronal origin of four of the monoclonal hybrid lines was shown by their immunoreactivity, following differentiation with 10 microM forskolin, to neurofilament protein, a neuron-specific marker. The monoclonal hybrid cell lines, but not the N18TG2 neuroblastoma, were shown to express an array of D1, D2, and D5 receptor mRNA as well as DARPP-32 mRNA. Two monoclonal cell lines expressed D1 receptor binding sites (X57, 29.2 +/- 4.5 fmol/mg protein and X62, 43.8 +/- 6.8 fmol/mg protein) which mediated the stimulation of adenylate cyclase activity. One cell line, X58, expressed only D2 dopamine receptors (80.9 +/- 9.8 fmol/mg protein) which were negatively coupled to adenylate cyclase activity. These findings suggest that the immortalized monoclonal hybrid cell lines are of neuronal origin and have incorporated elements of the medium spiny and cholinergic neurons of the developing striatum.