Short-term modulation at synapses between neurons in laminae II-V of the rodent spinal dorsal horn.

Unitary excitatory (EPSP) and inhibitory (IPSP) postsynaptic potentials (PSPs) were evoked between neurons in Rexed's laminae (L)II-V of spinal slices from young hamsters (7-24 days old) at 27°C using paired whole cell recordings. Laminar differences in synaptic efficacy were observed: excitatory connections were more secure than inhibitory connections in LII and inhibitory linkages in LII were less reliable than those in LIII-V. A majority of connections displayed paired-pulse facilitation or depression. Depression was observed for both EPSPs and IPSPs, but facilitation was seen almost exclusively for IPSPs. There were no frequency-dependent shifts between facilitation and depression. Synaptic depression was associated with an increased failure rate and decreased PSP half-width for a majority of connections. However, there were no consistent changes in failure rate or PSP time course at facilitating connections. IPSPs evoked at high-failure synapses had consistently smaller amplitude and showed greater facilitation than low-failure connections. Facilitation at inhibitory connections was positively correlated with synaptic jitter and associated with a decrease in latency. At many connections, the paired-pulse ratio varied from trial to trial and depended on the amplitude of the first PSP; dependence was greater for inhibitory synapses than excitatory synapses. Paired-pulse ratios for connections onto neurons with rapidly adapting, "phasic" discharge to depolarizing current injection were significantly greater than for connections onto neurons with tonic discharge properties. These results are evidence of diversity in synaptic transmission between dorsal horn neurons, the nature of which may depend on the types of linkage, laminar location, and intrinsic firing properties of postsynaptic cells.

The organization of projections from the olfactory bulb to the piriform cortex and olfactory tubercle in the rat.

The organization of the projection of olfactory bulb output cells was studied in the rat by injection of horseradish peroxidase (HRP) into the piriform cortex or olfactory tubercle. We made single HRP injections into small cuts in the fiber layer of the projection areas in order to enhance uptake by axons and to confine the region of HRP uptake. Following most of these injections, HRP-labeled axons could be traced in discrete fascicles through the fiber layer of the cortex or tubercle. These observations indicate that axons innervating the piriform cortex do not emit many long collaterals after they leave the lateral olfactory tract. HRP-labeled cells were generally observed throughout the ipsilateral olfactory bulb, but there were regions of greater density of labeled cells that differed in the various brains. The differences among the distributions of labeled mitral and tufted cells were analyzed statistically in 39 brains to test whether they varied systematically with injection site. In these analyses, the olfactory bulb was divided into 30 standard regions, and the labeled cells in each regions were counted. The distributions of labeled cells were similar for brains where injections were made into similar regions of the piriform cortex. The variations in density of labeled cells of the dorsal and anterior regions of the olfactory bulb were most strongly correlated with the positions of cortical injections. In contrast, the posterior medial regions of the bulb were heavily labeled after almost all injections. The ventral portions of the olfactory bulb were most heavily labeled after injections into the olfactory tubercle.

Synaptic connectivity of local circuit neurons in laminae III and IV of hamster spinal cord.

The present study was undertaken to examine the morphological bases of local synaptic interactions between dorsal horn interneurons. Seven interneurons responding to innocuous mechanical stimuli were intracellularly recorded in lamina III/IV of an isolated preparation of hamster spinal cord with partially intact innervation from an excised patch of hairy skin. Axonal arborizations were stained with horseradish peroxidase (HRP) and examined with an electron microscope. Five cells had extensive synaptic terminations (375-1,785 boutons/axon) with localized distributions (rostrocaudal distance, 425-1,251 microns) overlapping the dendritic trees. Two cells gave rise to deep stem axons that bifurcated into rostrocaudal daughter branches with collaterals ventral to the parent cell bodies (79-661 boutons/axon). Axons of local interneurons were thinly myelinated and formed terminal and en passant enlargements (mean [+/- S.D.] diameter = 0.88 +/- 0.24 microns, n = 157) containing clear, round vesicles 20-60 nm in diameter. Collateral branches of deep axon cells produced round, vesicle-containing boutons comparable in diameter (0.93 +/- 0.22 microns, n = 31) to local axon cells. Both types of interneurons formed asymmetric synaptic contacts with dendritic profiles, but not with cell bodies or axon terminals. Postsynaptic profiles contained sparse ribosomes and had a mean diameter of 1.0 +/- 0.5 microns (n = 49), significantly smaller than a population of identified proximal dendrites (2.3 +/- 0.9 microns, n = 47). HRP-labeled boutons were rarely (5/45 or 11%) in synaptic contact with more than one profile. We conclude that lamina III/IV interneurons make axodendritic synapses predominantly with distal dendrites. Thus, terminations of deep dorsal horn interneurons appear to have a postsynaptic distribution overlapping with axodendritic contacts formed by several functional classes of cutaneous sensory fibers signaling innocuous mechanical stimuli. Such overlap suggests that local spinal networks selectively and strongly influence afferent signals at initial stages of somatosensory integration.

Immunocytochemical localization of glutamic acid decarboxylase in physiologically identified interneurons of hamster spinal laminae III-V.

Neurons in Rexed's laminae III-V of an isolated spinal cord-skin patch preparation from hamsters were recorded in whole-cell mode and stained intracellularly with biocytin. Evidence of inhibitory synaptic function was obtained via post-hoc immunofluorescent labeling with a monoclonal antibody directed against an axon terminal isoform of brain glutamic acid decarboxylase. For a subset of neurons, examination with laser scanning confocal microscopy revealed punctate accumulations of glutamic acid decarboxylase immunoreactivity within axon enlargements (1-3 microm diameter), as imaged in single optical sections and confirmed by subsequent optical scans in the orthogonal plane. Axons of glutamic acid decarboxylase-immunoreactive neurons were found to exhibit dense local terminations overlapping the soma and dendrites or bifurcated into lengthy rostrocaudal daughter branches ventral to the cell body. The degree and uniformity of immunolabeling in axonal enlargements varied considerably, even amongst boutons belonging to the same cell. Glutamic acid decarboxylase-positive neurons received input from myelinated (A) afferent fibers and responded to natural stimuli appropriate for activating responses in low threshold mechanoreceptors. These results provide evidence that two different populations of GABAergic inhibitory interneurons are involved in local and intersegmental circuits that mediate integration of mechanosensory information in the deep spinal dorsal horn.

Selective suppression of endogenous peroxidase activity: application for enhancing appearance of HRP-labeled neurons in vitro.

A simple procedure to suppress selectively the endogenous peroxidase activity of red blood cells in histological sections of the mammalian nervous system is described. Pretreatment of sections with a series of ethanol solutions results in selective abolition of red blood cell staining and also leads to enhanced visualization of neurons that have been injected with horseradish peroxidase (HRP). The method is particularly useful for processing in vitro brain slices that contain HRP-labeled neurons. It can also be used for processing unperfused neural tissue from in vivo HRP labeling experiments. The ethanol pretreatment is compatible with several standard histochemical techniques for the demonstration of HRP.

Slow potentials activated by afferent and direct stimulation in spinal interneurons of laminae III-V.

Stimulation of hairy skin afferent fibers and/or direct depolarizing current injection initiates persistent changes in membrane potential (slow potentials), lasting up to 1 min or longer, in some deep dorsal horn neurons. Intracellular staining with horseradish peroxidase shows that these cells are interneurons, suggesting that slow potentials modulate moment-to-moment impulse transmission in local spinal circuits.

Demonstration of artifactual coupling between spinal neurons and glial cells during intracellular recording with micropipette electrodes.

Intracellular recordings with electrophysiological properties that are characteristic of both neurons and glial cells were obtained in the isolated spinal dorsal horn. Intracellular staining supports an idea that these elements may become artifactually coupled via the recording electrode. This coupling can be a gradual process, occurring over periods up to 30 min, and may have implications for the interpretation of slow, afferent-evoked potentials in some dorsal horn neurons.

Selective excitation of neurons in the mammalian spinal dorsal horn by aspartate and glutamate in vitro: correlation with location and excitatory input.

The electrical activity of mammalian dorsal horn neurons was recorded with pipette microelectrodes in an in vitro spinal cord slice preparation with dorsal roots intact. Addition of relatively low concentrations of aspartate or glutamate to the superfusion solution or through the recording pipette with small iontophoretic currents excited only a subset of neurons. The majority of these excited neurons were located in the superficial dorsal horn (Rexed's laminae I and II) and a preponderance were excited by the C-fiber components of dorsal root volleys. These findings are consistent with the idea that aspartate or glutamate may function as a synaptic mediator for some neurons terminating in the superficial dorsal horn.

Effects of one-stage or serial transections of the lateral olfactory tracts on behavior and plasma testosterone levels in male hamsters.

Bilateral transections of the later olfactory (LOT) in male hamsters at the rostral border of the olfactory tubercle or anterior to the tubercle produced severe deficits in mounting, scent marking, food hoarding and nesting behaviors. There were no impairments of these behaviors following unilateral transections. Cutting of the LOT in two stages, and provision of additional experience in sexual and territorial behaviors during the interval between the first and second transection, did not alleviate the behavioral deficits. Impairments in mounting and scent marking were dissociated in some subjects, indicating individual differences in the hamsters' dependence on chemosensory cues for the regulation of these two types of social behavior. It is suggested that the apparent sparing of behavioral function which has been associated with gradual destruction of nervous tissue is due to the development of alternate mechanisms of response to environmental cues, and may be more difficult when brain damage is in structures which initially process these cues. Although sexual and territorial behaviors in the hamster are dependent on gonadal hormones, the observed impairments of these behaviors could not be explained as indirect consequences of lesion effects on the hypophysiogonadal axis. Circulating testosterone (T) levels in experimental subjects were not different from levels in control (sham-operated) subjects. An inverse correlation was observed between the control subjects' circulating T levels and their incidence of scent marking. A similar correlation was observed between postoperative T levels and preoperative marking rates in experimental subjects, despite the fact that most subjects failed to scent mark after bilateral LOT transections. The latter observation strengthens the inference that the bilateral deficits were independent of changes in basal hormone levels.

Dispersive analysis of the pion transition form factor.

We analyze the pion transition form factor using dispersion theory. We calculate the singly-virtual form factor in the time-like region based on data for the [Formula: see text] cross section, generalizing previous studies on [Formula: see text] decays and [Formula: see text] scattering, and verify our result by comparing to [Formula: see text] data. We perform the analytic continuation to the space-like region, predicting the poorly-constrained space-like transition form factor below [Formula: see text], and extract the slope of the form factor at vanishing momentum transfer [Formula: see text]. We derive the dispersive formalism necessary for the extension of these results to the doubly-virtual case, as required for the pion-pole contribution to hadronic light-by-light scattering in the anomalous magnetic moment of the muon.

Visceral hypersensitivity in female but not in male serotonin transporter knockout rats.

BACKGROUND: Visceral hypersensitivity occurs in irritable bowel syndrome (IBS), particularly in women. Serotonin signaling, including reduced serotonin transporter (SERT) expression, may be disrupted in IBS patients. We studied SERT gene knockout (KO) rats to determine if they exhibited sex-related alterations in visceral sensitivity. METHODS: We measured serotonin in the colonic mucosa using HPLC and amperometric microelectrode techniques. Visceral sensitivity was assessed using the electromyographic visceromotor response (VMR) in response to colorectal balloon distention (CRD). We studied the electrophysiologic properties of colon projecting sensory neurons in vitro using whole-cell recordings. KEY RESULTS: Mucosal serotonin levels were not different among male and female WT and SERT KO rats. Serotonin oxidation currents in vitro were larger (P < 0.05) in tissues from male and female SERT KO compared with WT rats. Oxidation currents in male and female WT, but not SERT KO, rats were increased (P < 0.05) by the SERT inhibitor fluoxetine (1 µmol L(-1) ). The VMR to CRD was increased in female but not in male SERT KO rats (P < 0.05); this response varied with the estrous cycle. Colon projecting sensory neurons from female SERT KO rats fired more action potentials compared with neurons from female WT rats. There were no differences in action potential firing in neurons from male WT and SERT KO rats. CONCLUSIONS & INFERENCES: Increased colonic extracellular serotonin in female SERT KO rats is associated with visceral hypersensitivity and hyperexcitability of colon projecting sensory neurons. The SERT KO rat is a model for studying interactions between serotonin, sex and visceral sensation.

Musculoskeletal injuries in construction: a review of the literature.

The first step in addressing any problem is recognition of the problem and a measure of its size and scope. There have been few reviews to date of the evidence of a musculoskeletal disorders problem in construction, particularly in the United States. Construction contractors in the United States have questioned the existence of a musculoskeletal disorders problem in construction, so a review of the evidence is warranted. The types of evidence reviewed include: 1) historical evidence, 2) injury data, 3) workers' compensation data, 4) medical exam data, 5) survey data, and 6) exposure data. Injury data generally represent injuries that the employers have identified as work-related and recorded or reported. Workers' compensation data are from cases that have been filed by workers for compensation and quite often represent only "closed" cases where compensation has been awarded. Medical exam data are from physical examinations of workers. Symptom survey data are the most inclusive and show the number of workers who self-report musculoskeletal problems. Exposure data include measurements made of exposure to musculoskeletal risk factors. The existing data show construction workers to be at significant risk of musculoskeletal injury, specifically related to the work they do. Their risk of musculoskeletal injury is much higher than that of other workers who have less heavy work, about 50 percent higher than all other workers. Several trades have been extensively studied, while others have been studied to a lesser extent. While the exact relationship between exposures and injuries is complex and often multifaceted, it would be difficult to deny the existence of the problem and the fact that these injuries are, to a great extent, related to the work that construction workers perform.

Functional properties and axon terminations of interneurons in laminae III-V of the mammalian spinal dorsal horn in vitro.

1. The functional organization of interneurons in spinal laminae III-V was studied in an isolated preparation of hamster dorsal horn with sensory innervation from an excised skin patch. Morphological details of 40 neurons were visualized by intracellular injection of horseradish peroxidase. Active and passive membrane properties, synaptic responses to cutaneous nerve volleys, and responses to innocuous mechanical stimuli were determined for 25 cells with identified axons. 2. Neurons were classified into two types: 1) cells with local axons, branching in proximity to the cell soma and dendrites, that produced numerous synaptic boutons (740 +/- 504/axon; mean +/- SD), often arranged in clusters and 2) neurons with deep axons that usually bifurcated into rostral and caudal daughter branches up to 2.5 mm long, giving off collaterals ventral to the cell body and dendrites and forming significantly fewer boutons (155 +/- 140/axon) than local axon cells. A majority of boutons of local axon and deep axon cells, 89 and 83%, respectively, were of the en passant type. 3. Dendritic trees of local axon cells were relatively compact dorsoventrally (119 +/- 42 microns) and mediolaterally (128 +/- 45 microns), but were elongated rostrocaudally (404 +/- 121 microns). In comparison, dendritic trees of deep axon cells radiated significantly farther dorsoventrally (218 +/- 88 microns) and mediolaterally (180 +/- 34 microns), but exhibited comparable rostrocaudal spread (413 +/- 128 microns). There was no correlation between dorsoventral and mediolateral dendritic spread and mediolateral soma location for either cell type. However, for medially situated deep axon cells the rostrocaudal dendritic spread was up to 180% greater than for those located laterally. For nearly one-half of all cells (49%; 17/35) dendritic processes extended dorsally into lamina II. 4. Local axon cells had resting membrane potentials that were more negative than deep axon cells (-59.5 +/- 6.1 and -53.6 +/- 4.7 mV, respectively), but the amplitude and duration of action potentials generated by the two types were similar. Neuronal input resistance (RN) and membrane time constant (tau m) varied widely from cell to cell, but were not significantly different for local axon (77.4 +/- 46.8 M omega, 13.4 +/- 9.5 ms) and deep axon cells (46.5 +/- 19.2 M omega, 6.6 +/- 3.0 ms). 5. Volleys in myelinated afferent fibers activated fast rising excitatory postsynaptic potentials (EPSPs) that exhibited later, more slowly rising potentials with multiple components in a majority of deep axon (89%) and local axon (72%) neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

Spike frequency adaptation and signaling properties of identified neurons in rodent deep spinal dorsal horn.

Using whole cell recordings, I analyzed the intrinsic discharge properties for 285 neurons in Rexed's laminae III-V of isolated hamster spinal cord preparations. Neurons were characterized by their responses to step-wise and ramp-hold depolarizing current applied through the recording pipettes. Tonic cells (133/285; 47%) fired repetitively during step-wise current application. Firing decayed linearly (-0.14 to -4.3 imp . s(-1) . s(-1)) or was bimodal, with an initial exponential phase (tau approximately 450 ms) followed by a linear decline (-0.02 to -6.3 imp . s(-1) . s(-1)); discharge frequency was unrelated to current trajectory. Phasic-firing cells (108/285; 38%) responded with a burst discharge having an initial rapid, exponential decrease (tau approximately 30 ms) and subsequent linear decline (-1 to -78 imp . s(-1) . s(-1)). Phasic cells were activated preferentially by fast current ramps (slope, 70 pA/s-2.2 nA/s) with the number and frequency of impulses increasing with current slope. Delayed-firing cells (44/285; 15%), responded to current steps with an accelerating firing following a substantial latent period (0.5-4 s) and discharged during current ramps with slopes less than approximately 100 pA/s. Intracellular staining revealed a significant association between electrophysiological profile and neuronal morphology. A majority of presumed projection cells (22/30; 73%) exhibited tonic firing to step-wise activation. The preponderance of phasic and delayed firing cells, 93% (42/45) and 71% (12/17), respectively, were interneurons with local or intersegmental terminations. Differential sensitivity to static and time-varying components of membrane current suggest differences in neuronal signaling properties that may have important implications for integration of mechanosensory information in the deep spinal dorsal horn.

Synaptic mediation from cutaneous mechanical nociceptors.

1. Responses of dorsal horn neurons to cutaneous mechanical stimulation were studied in an in vitro preparation of hamster spinal cord with partially intact innervation from an isolated patch of hairy skin. Stable extracellular and intracellular recordings were obtained from cells with different mechanoreceptive properties similar to those reported for other species in vivo. Analyses were made of the intracellular responses of 25 dorsal horn neurons activated selectively by mechanical stimulation to the skin patch. 2. Bath application of the broad spectrum, excitatory amino acid (EAA) receptor antagonist, kynurenic acid (1 mM) blocked excitation of 7 of 8 high-threshold mechanoreceptive units by either cutaneous nerve volleys or mechanical stimulation of the skin. This concentration of kynurenic acid suppressed peripherally evoked responses in 8 of 14 neurons responsive to innocuous mechanical stimuli. 3. High-threshold mechanoreceptive neurons of the superficial dorsal horn exhibited one of three distinctive patterns of postsynaptic potentials in response to electrical stimulation of cutaneous afferent fibers: 1) a simple fast excitatory postsynaptic potential (EPSP), 2) a fast EPSP with a prolonged decay phase lasting between 100 and 1,000 ms, and 3) a multiphasic response dissociable on the basis of stimulus strength consisting of a fast EPSP followed by a hyperpolarizing inhibitory postsynaptic potential (IPSP) (duration 80-100 ms). Gentle mechanical stimuli initiated inhibition from areas adjacent to the high-threshold mechanically excitatory field; this suggests that membrane hyperpolarization in these neurons was evoked by input from low-threshold mechanoreceptors. 4. Bath application of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM), a competitive EAA antagonist selective for non-N-methyl-D-aspartate (non-NMDA) receptor subtypes, substantially or completely (56-100%) suppressed EPSPs evoked from cutaneous afferent fibers in high-threshold mechanoreceptive neurons. CNQX also decreased the membrane depolarization, the frequency of EPSPs, and the frequency of action potentials evoked by mechanical stimulation of the receptive field. 5. CNQX (10 microM) or kynurenic acid (1 mM) had considerably weaker effects on IPSPs than on EPSPs evoked from the periphery in superficial dorsal horn neurons. IPSP amplitudes were unchanged by these agents in some neurons and decreased by only 20-25% in others. 6. We conclude that L-glutamate acting on non-NMDA receptors mediates fast synaptic excitation of superficial dorsal horn neurons from peripheral mechanical nociceptors with myelinated fibers. Furthermore, the observations imply either an agent other than L-glutamate or one acting at different membrane receptors is a synaptic mediator for other peripheral afferent units including some activated by innocuous mechanical stimuli.

Involvement of GABA and glycine in recurrent inhibition of spinal motoneurons.

1. Recurrent inhibitory postsynaptic potentials (IPSPs) were recorded intracellularly from chloride-loaded motoneurons in the isolated lumbar spinal cord of neonatal rats (day 5-day 12). This in vitro preparation exhibited an intact and functional recurrent inhibitory pathway that displayed characteristics previously described for this pathway in other species. 2. Although strychnine (1-5 microM) depressed the chloride-dependent recurrent synaptic potentials evoked by ventral root stimulation by 48.2 +/- 2.7% (mean +/- SE, n = 13), confirming that part of the recurrent IPSP is mediated by a glycinergic mechanism, in every case a residual strychnine-resistant synaptic potential was observed. 3. The gamma-aminobutyric acid (GABA) antagonist bicuculline, in low concentrations (2-10 microM), depressed the recurrent synaptic potentials in a dose-dependent manner by 27.0 +/- 4.3% (range 0-49%, n = 19). Application of bicuculline almost eliminated the strychnine-resistant component of the IPSP. However, in some motoneurons, a small synaptic potential remained after combined application of strychnine and bicuculline. 4. The selective antagonists of GABA uptake, (+/-)-nipecotic acid (1 mM) and guvacine (1 mM), increased the amplitude of recurrent synaptic potentials in 12 of 16 motoneurons by 37.2 +/- 7.2% (range 12.6-84.2%). 5. The excitatory amino acid antagonists kynurenic acid (1 mM), 6-cyano-7-nitroquinoxaline-2,3-dione [CNQX (10 microM)] and 6,7-dinitroquinoxaline-2,3-dione (10 microM) potentiated recurrent synaptic potentials in 5 of 7 motoneurons. However, CNQX (10-15 microM) in the presence of strychnine and bicuculline virtually abolished the synaptic potential remaining after application of the inhibitory amino acid antagonists. It is concluded that ventral root stimulation evokes a small excitatory amino acid-mediated synaptic potential in neonatal rat motoneurons. 6. An antidromic synaptic potential due to electrotonic coupling between motoneurons was unaffected by changes in membrane potential, chloride loading, or antagonists of glycine, GABA, excitatory amino acid, and acetylcholine receptors. 7. The results suggest that a major portion of the strychnine-resistant component of the IPSP is mediated by a GABAergic mechanism. It is concluded that both glycinergic and GABAergic mechanisms play a role in recurrent inhibition of motoneurons in the mammalian spinal cord. It is unknown whether these inhibitory amino acids are released by a single pool of Renshaw cells or by neurochemically distinct populations.

Comparison of primary afferent and glutamate excitation of neurons in the mammalian spinal dorsal horn.

The actions of L-glutamate and agonists, agents blocking their membrane receptors and dorsal root afferent volleys, were compared on intracellularly recorded neuronal activity in an in vitro horizontal slice preparation of the hamster spinal dorsal horn. Bath-applied L-glutamate or L-aspartate (less than or equal to 1 mM) rapidly depolarized and excited less than a third of the dorsal horn neurons sampled. Bathing solutions containing low Ca2+ eliminated synaptic transmission in the slices but failed to block the excitatory effects of L-glutamate for the majority of the neurons tested. N-Acetylaspartylglutamate had no effect on dorsal horn neurons at concentrations up to 1 mM. Neurons excited by L-glutamate were most commonly located in the superficial dorsal horn (laminae I and II). Neurons insensitive to L-glutamate were more broadly distributed, with a number being located in laminae III-V. Kynurenic acid, 2-amino-4-phosphonobutyric acid, and 2,3-piperidine dicarboxylic acid selectively antagonized rapid, short-lasting synaptic components of the dorsal cord potentials. Kynurenic acid reversibly antagonized intracellularly recorded L-glutamate-induced excitation, spontaneous synaptic potentials, and fast synaptic potentials evoked by dorsal root volleys. Compounds with strong antagonist actions at the NMDA receptor, 2-amino-5-phosphonovaleric acid and D-alpha-aminoadipic acid, were much less effective in suppressing the effects of L-glutamate or in blocking synaptic potentials. We conclude that a subset of spinal neurons directly excited by dorsal root fibers have excitatory membrane receptors activated by L-glutamate. This conclusion is consistent with the concept that L-glutamate or a substance binding to the receptors it activates is released from the central terminals of some primary afferent fibers and mediates fast synaptic transmission from them to certain spinal neurons in the dorsal horn.