Functional Properties of Sensory Nerve Terminals of the Mouse Cornea.

Purpose: To define the firing properties of sensory nerve terminals innervating the adult mouse cornea in response to external stimuli of differing modality. Methods: Extracellular electrical activity of single corneal sensory nerve terminals was recorded in excised eyes of C57BL/6J mice. Eyes were placed in a recording chamber and were continuously superfused with warm saline solution. Nerve terminal impulse (NTI) activity was recorded by means of a glass pipette (tip ∼ 50 µm), applied on the corneal surface. Nerve terminal impulse discharges were stored in a computer for offline analysis. Results: Three functionally distinct populations of nerve terminals were identified in the mouse cornea. Pure mechanonociceptor terminals (9.5%) responded phasically and only to mechanical stimuli. Polymodal nociceptor terminals (41.1%) were tonically activated by heat and hyperosmolal solutions (850 mOsm·kg-1), mechanical force, and/or TRPV1 and TRPA1 agonists (capsaicin and allyl isothiocyanate [AITC], respectively). Cold-sensitive terminals (49.4%) responded to cooling. Approximately two-thirds of them fired continuously at 34°C and responded vigorously to small temperature reductions, being classified as high-background activity, low-threshold (HB-LT) cold thermoreceptor terminals. The remaining one-third exhibited very low ongoing activity at 34°C and responded weakly to intense cooling, being named low-background activity, high-threshold (LB-HT) cold thermoreceptor terminals. Conclusions: The mouse cornea is innervated by trigeminal ganglion (TG) neurons that respond to the same stimulus modalities as corneal receptors of other mammalian species. Mechano- and polymodal endings underlie detection of mechanical and chemical noxious stimuli while HB-LT and LB-HT cold thermoreceptors appear to be responsible for basal and irritation-evoked tearing and blinking, respectively.

Deletion of the murine ATP/UTP receptor P2Y2 alters mechanical and thermal response properties in polymodal cutaneous afferents.

P2Y2 is a member of the P2Y family of G protein-coupled nucleotide receptors that is widely co-expressed with TRPV1 in peripheral sensory neurons of the dorsal root ganglia. To characterize P2Y2 function in cutaneous afferents, intracellular recordings from mouse sensory neurons were made using an ex vivo preparation in which hindlimb skin, saphenous nerve, dorsal root ganglia and spinal cord are dissected intact. The peripheral response properties of individual cutaneous C-fibers were analyzed using digitally controlled mechanical and thermal stimuli in male P2Y2(+/+) and P2Y2(-/-) mice. Selected sensory neurons were labeled with Neurobiotin and further characterized by immunohistochemistry. In wildtype preparations, C-fibers responding to both mechanical and thermal stimuli (CMH or CMHC) preferentially bound the lectin marker IB4 and were always immunonegative for TRPV1. Conversely, cells that fired robustly to noxious heat, but were insensitive to mechanical stimuli, were TRPV1-positive and IB4-negative. P2Y2 gene deletion resulted in reduced firing by TRPV1-negative CMH fibers to a range of heat stimuli. However, we also identified an atypical population of IB4-negative, TRPV1-positive CMH fibers. Compared to wildtype CMH fibers, these TRPV1-positive neurons exhibited lower firing rates in response to mechanical stimulation, but had increased firing to noxious heat (43-51°C). Collectively, these results demonstrate that P2Y2 contributes to response properties of cutaneous afferents, as P2Y2 deletion reduces responsiveness of conventional unmyelinated polymodal afferents to heat and appears to result in the acquisition of mechanical responsiveness in a subset of TRPV1-expressing afferents.

Arsenite induces neurotoxic effects on AFD neurons via oxidative stress in Caenorhabditis elegans.

Arsenic permeates our environment. As a result, humans are continually exposed to it. This study investigates the possible roles of oxidative stress in arsenite (As(III))-induced neurotoxicity in Caenorhabditis elegans. Exposure to As(III), at the concentrations examined, caused a decrease in locomotor behaviors (frequencies of body bends, head thrashes, and reversals) of C. elegans. In addition, As(III) exposure (100 µM) decreased thermotactic behaviors, and induced severe deficits in the structural properties of AFD sensory neurons. Exposure to As(III) (100 µM) also caused an elevated production of intracellular reactive oxygen species (ROS) in wild-type C. elegans. Pretreatment with the antioxidant curcumin ameliorated the decrease in locomotor and thermotactic behavior, the formation of deficits in the structural properties of AFD sensory neurons, and intracellular ROS in As(III)-exposed nematodes. Our study suggests that oxidative stress plays a crucial role in the As(III)-induced neurotoxic effects on locomotor behavior and the structures and function of AFD sensory neurons in As(III)-exposed nematodes.

A gustatory receptor paralogue controls rapid warmth avoidance in Drosophila.

Behavioural responses to temperature are critical for survival, and animals from insects to humans show strong preferences for specific temperatures. Preferred temperature selection promotes avoidance of adverse thermal environments in the short term and maintenance of optimal body temperatures over the long term, but its molecular and cellular basis is largely unknown. Recent studies have generated conflicting views of thermal preference in Drosophila, attributing importance to either internal or peripheral warmth sensors. Here we reconcile these views by showing that thermal preference is not a singular response, but involves multiple systems relevant in different contexts. We found previously that the transient receptor potential channel TRPA1 acts internally to control the slowly developing preference response of flies exposed to a shallow thermal gradient. We now find that the rapid response of flies exposed to a steep warmth gradient does not require TRPA1; rather, the gustatory receptor GR28B(D) drives this behaviour through peripheral thermosensors. Gustatory receptors are a large gene family, widely studied in insect gustation and olfaction, and are implicated in host-seeking by insect disease vectors, but have not previously been implicated in thermosensation. At the molecular level, GR28B(D) misexpression confers thermosensitivity upon diverse cell types, suggesting that it is a warmth sensor. These data reveal a new type of thermosensory molecule and uncover a functional distinction between peripheral and internal warmth sensors in this tiny ectotherm reminiscent of thermoregulatory systems in larger, endothermic animals. The use of multiple, distinct molecules to respond to a given temperature, as observed here, may facilitate independent tuning of an animal's distinct thermosensory responses.

The evaporative function of cockroach hygroreceptors.

Insect hygroreceptors associate as antagonistic pairs of a moist cell and a dry cell together with a cold cell in small cuticular sensilla on the antennae. The mechanisms by which the atmospheric humidity stimulates the hygroreceptive cells remain elusive. Three models for humidity transduction have been proposed in which hygroreceptors operate either as mechanical hygrometers, evaporation detectors or psychrometers. Mechanical hygrometers are assumed to respond to the relative humidity, evaporation detectors to the saturation deficit and psychrometers to the temperature depression (the difference between wet-bulb and dry-bulb temperatures). The models refer to different ways of expressing humidity. This also means, however, that at different temperatures these different types of hygroreceptors indicate very different humidity conditions. The present study tested the adequacy of the three models on the cockroach's moist and dry cells by determining whether the specific predictions about the temperature-dependence of the humidity responses are indeed observed. While in previous studies stimulation consisted of rapid step-like humidity changes, here we changed humidity slowly and continuously up and down in a sinusoidal fashion. The low rates of change made it possible to measure instantaneous humidity values based on UV-absorption and to assign these values to the hygroreceptive sensillum. The moist cell fitted neither the mechanical hygrometer nor the evaporation detector model: the temperature dependence of its humidity responses could not be attributed to relative humidity or to saturation deficit, respectively. The psychrometer model, however, was verified by the close relationships of the moist cell's response with the wet-bulb temperature and the dry cell's response with the dry-bulb temperature. Thus, the hygroreceptors respond to evaporation and the resulting cooling due to the wetness or dryness of the air. The drier the ambient air (absolutely) and the higher the temperature, the greater the evaporative temperature depression and the power to desiccate.

Regulation of TRPM8 channel activity.

Transient Receptor Potential Melastatin 8 (TRPM8) is a Ca(2+) permeable non-selective cation channel directly activated by cold temperatures and chemical agonists such as menthol. It is a well established sensor of environmental cold temperatures, found in peripheral sensory neurons, where its activation evokes depolarization and action potentials. The activity of TRPM8 is regulated by a number of cellular signaling pathways, most notably by phosphoinositides and the activation of phospholipase C. This review will summarize current knowledge on the physiological and pathophysiological roles of TRPM8 and its regulation by various intracellular messenger molecules and signaling pathways.

Distribution of trigeminothalamic and spinothalamic lamina I terminations in the cat.

The distribution in the thalamus of terminal projections from lamina I neurons of the trigeminal, cervical, and lumbosacral dorsal horn was investigated with the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) in the cat. Iontophoretic injections were guided by single- and multi-unit physiological recordings. The injections in particular cases were essentially restricted to lamina I, whereas in others they spread across laminae I-III or laminae I-V. The trigemino- and spinothalamic (TSTT) terminations were identified immunohistochemically. In all cases, regardless of the level of the injections, terminal fibers were consistently distributed in three main locations: the submedial nucleus; the ventral aspect of the basal ventral medial nucleus and ventral posterior nuclei; and, the dorsomedial aspect of the ventral posterior medial nucleus. The terminal fields in the submedial nucleus and the ventral aspect of the ventral posterior group were topographically organized. Terminations along the ventral aspect of the ventral posterior group extended posterolaterally into the caudal part of the posterior nucleus and anteromedially into the ventromedial part of the ventral lateral nucleus. In several cases with trigeminal lamina I injections, a terminal labeling patch was observed within the core of the ventral posterior medial nucleus. In cases with spinal lamina I injections, terminations were also consistently found in the lateral habenula, the parafascicular nucleus, and the nucleus reuniens. Isolated terminal fibers were occasionally seen in the zona incerta, the dorsomedial hypothalamus, and other locations. These anatomical observations extend prior studies of TSTT projections and identify lamina I projection targets that are important for nociceptive, thermoreceptive, and homeostatic processing in the cat. The findings are consistent with evidence from physiological (single-unit and antidromic mapping) and behavioral studies. The novel identification of spinal lamina I input to the lateral habenula could be significant for homeostatic behaviors.

Identification and function of thermosensory neurons in Drosophila larvae.

Although the ability to sense temperature is critical for many organisms, the underlying mechanisms are poorly understood. Using the calcium reporter yellow cameleon 2.1 and electrophysiological recordings, we identified thermosensitive neurons and examined their physiologic response in Drosophila melanogaster larvae. In the head, terminal sensory organ neurons showed increased activity in response to cooling by < or =1 degrees C, heating reduced their basal activity, and different units showed distinct response patterns. Neither cooling nor heating affected dorsal organ neurons. Body wall neurons showed a variety of distinct response patterns to both heating and cooling; the diverse thermal responses were strikingly similar to those described in mammals. These data establish a functional map of thermoresponsive neurons in Drosophila larvae and provide a foundation for understanding mechanisms of thermoreception in both insects and mammals.

Projection neurons originating from thermo- and hygrosensory glomeruli in the antennal lobe of the cockroach.

Most insects are equipped with specialized thermo- and hygroreceptors to locate a permissible range of ambient temperature and distant water sources, respectively. In the cockroach, Periplaneta americana, cold, moist, and dry receptor cells in the antennae send axons to particular sets of two or three glomeruli in the dorsocentral part of the antennal lobe (primary olfactory center), designated DC1-3 glomeruli. However, it is not known how thermo- and hygrosensory signals from these glomeruli are represented in higher-order centers, the protocerebrum, in any insect species. With the use of intracellular recording and staining techniques, we identified a new class of interneurons with dendrites almost exclusively in the DC1, DC2, or DC3 glomeruli and axons projecting to the protocerebrum in the cockroach. Remarkably, terminals of all these projection neurons (PNs) covered almost identical areas in the lateral protocerebrum (LP), although their termination areas outside the LP differed from neuron to neuron. The termination areas within the LP were distinct from, but close to, those of uniglomerular and macroglomerular PNs that transmitted signals concerning general odors and female sex pheromones, respectively. PNs originating from DC1, DC2, and DC3 glomeruli exhibited excitatory responses to cold, moist, and dry stimuli, respectively, probably due to excitatory synaptic input from cold, moist, and dry receptor cells, respectively, whereas their responses were often modulated by olfactory stimuli. These findings suggested that dorsocentral PNs participate in neural pathways that lead to behavioral responses to temperature or humidity changes.

Specificity of cold thermotransduction is determined by differential ionic channel expression.

Sensations of cold are mediated by specific thermoreceptor nerve endings excited by low temperature and menthol. Here we identify a population of cold-sensitive cultured mouse trigeminal ganglion neurons with a unique set of biophysical properties. Their impulse activity during cooling and menthol application was similar to that of cold thermoreceptor fibers in vivo. We show that cooling closes a background K+ channel, causing depolarization and firing that is limited by the slower reduction of a cationic inward current (Ih). In cold-insensitive neurons, firing is prevented by a slow, transient, 4-AP-sensitive K+ current (IKD) that acts as an excitability brake. In addition, pharmacological blockade of IKD induced thermosensitivity in cold-insensitive neurons, a finding that may explain cold allodynia in neuropathic pain. These results suggest that cold sensitivity is not associated to a specific transduction molecule but instead results from a favorable blend of ionic channels expressed in a small subset of sensory neurons.

Vanilloid receptor 1-like receptor-immunoreactive primary sensory neurons in the rat trigeminal nervous system.

Immunohistochemistry for vanilloid receptor 1-like receptor (VRL-1), a candidate transducer for high-threshold noxious heat, was performed on rat trigeminal primary sensory neurons. The immunoreactivity was detected in 14% of the trigeminal ganglion cell bodies, while the neurons in the mesencephalic trigeminal tract nucleus were almost devoid of it (0.5%). The immunoreactive neurons in the trigeminal ganglion were mostly of medium to large size (mean+/-S.D. of 956+/-376microm(2)). Nerve bundles in the tooth pulp, periodontal ligament, facial skin and oral mucosa contained VRL-1-positive smooth nerve fibers. The immunoreactivity could not be traced to the isolated nerve fibers, except in the tooth pulp. In the brainstem trigeminal nuclear complex, a notable concentration of the immunoreactivity was seen in laminae I and II of the medullary dorsal horn. Thirty-seven per cent of the trigeminal ganglion neurons retrogradely labeled from the tooth pulp exhibited VRL-1 immunoreactivity, while the immunoreactivity was detected in only 9% of those labeled from the skin. Co-expression of calcitonin gene-related peptide was common among the VRL-1-immunoreactive tooth pulp neurons (45%) and cutaneous neurons (25%). Moreover, as many as 41% of the VRL-1-immunoreactive tooth pulp neurons co-expressed parvalbumin immunoreactivity. Parvalbumin immunoreactivity was never detected in the VRL-1-immunoreactive cutaneous neurons. From the findings of the present study, we propose that large primary neurons responding to high-threshold noxious heat are abundant in the tooth pulp, but not in the facial skin.

Behavioral state-dependent thermal feedback influencing the hypothalamic thermostat.

The experimental evidence on the behavioral state-dependent compartmentalization of temperature in the central nervous system of three homeothermic species has been reviewed to address the question of how selective brain cooling influences hypothalamic temperature regulation.

Neural elements in the human temporomandibular articular disc.

The purpose of this study was to determine the presence and describe the location of neural elements in the articular disc of the human temporomandibular joint. Six articular discs were obtained from three adult human subjects at autopsy. Four discs were cut into segments of known anterior-posterior orientation. The remaining two were processed intact. All tissues were stained in bulk with gold chloride, and frozen, sectioned on a sliding microtome at 70 to 100 microns, mounted on slides, dehydrated, and coverslipped. Nerve fibers were seen penetrating the discs from the pericapsular connective tissue. Structures resembling Ruffini endings, Pacinian corpuscles, and Golgi tendon organs were identified in the pericapsular connective tissue and within the disc. The population density of neural elements was the greatest at the periphery of the disc and progressively decreased towards the center, which was essentially devoid of them. The concentration of neural elements appeared to be greater at the anterior and posterior margins of the disc, with the greatest concentration being posteriorly. These findings support the theory that afferent nerves may arise from neural elements within the disc.

Anatomic evidence of nociceptive inputs to primary somatosensory cortex: relationship between spinothalamic terminals and thalamocortical cells in squirrel monkeys.

This study examined anatomic pathways that are likely to transmit noxious and thermal cutaneous information to the primary somatosensory cortex. Anterograde and retrograde labeling techniques were combined to investigate the relationship between spinothalamic (STT) projections and thalamocortical neurons in the squirrel monkey (Saimiri sciureus). Large injections of diamidino yellow (DY) were placed in the physiologically defined hand region of primary somatosensory cortex (hSI), and wheat germ agglutinin-horseradish peroxidase (WGA-HRP) was injected in the contralateral cervical enlargement (C5-T1). Both DY-labeled neuronal cell bodies and HRP-labeled STT terminal-like structures were visualized within single thalamic sections in each animal. Quantitative analysis of the positions and numbers of retrogradely labeled neurons and anterogradely labeled terminal fields reveal that: 1) ventral posterior lateral (VPL), ventral posterior inferior (VPI), and central lateral (CL), combined, receive 87% of spinothalamic inputs originating from the cervical enlargement; 2) these three nuclei contain over 91% of all thalamocortical neurons projecting to hSI that are likely to receive STT input; and 3) these putatively contacted neurons account for less than 24% of all thalamic projections to hSI. These results suggest that three distinct spinothalamocortical pathways are capable of relaying nociceptive information to the hand somatosensory cortex. Moreover, only a small portion of thalamocortical neurons are capable of relaying STT-derived nociceptive and thermal information to the primary somatosensory cortex.

[The morphology of sensory corpuscles in the joint capsule--ultrastructural and histochemical study].

Sensory corpuscles in the cat elbow joint were observed by light and electron microscopy. Two types of corpuscles were identified, i.e. Pacini-type and Ruffini-type corpuscles. Silver impregnation revealed a single straight axon terminal in Pacini-type and highly-ramified fine axons in Ruffini-type corpuscles. Pacini-type corpuscles, 100-200 microns long and 30 microns wide, consisted of dense lamellae of lamellar cells and thick capsule. The lamellar portion contained a centrally-located axon terminal, and was similar in organization to the inner core of the typical Pacini corpuscle. The capsule was the continuation of perineurial sheath. This corpuscle was devoid of outer core structure as seen in a typical Pacini corpuscle. Ruffini-type corpuscles, 50-150 microns long and 25-50 microns wide, had the branched axon terminals with varicosities under the incomplete capsule. Axons, which were surrounded by thin Schwann cell processes, were embedded in the dense layers of collagen fibrils. The interior of the corpuscle was separated into small compartments by cell processes extended from the capsule. The axon varicosities contained numerous mitochondria. These fine structures of the corpuscles were similar to those of Ruffini corpuscles reported so far in other regions. Both Pacini-type and Ruffini-type corpuscles were clearly demonstrated by histochemistry for ChE. Inasmuch as the staining feature of corpuscles was different from each other, the distribution pattern of corpuscles in the joint capsules could be obtained by examining semi-serial sections. Pacini-type corpuscles were mainly found in the synovial layer, while Ruffini-type corpuscles were mainly located in the fibrous layer of the joint capsule. Both types of corpuscles were located near the median nerve on the flexor side of the joint. The reaction products of ChE activity were located in the peri-axonal as well as inter-lamellar spaces of Pacini-type corpuscles, and in the peri-axonal region as well as around Schwann cell processes of Ruffini-type corpuscles. No definite reaction product was found within axon terminals. Some reaction products were also found in the rough endoplasmic reticulum and/or nuclear envelope, suggesting that ChE is synthesized by Schwann cells. The significance of these sensory corpuscles with regard to the deep sensation in the joint capsule was discussed from the point of view of the electro-physiological characteristics of the corpuscles.