Confocal imaging of HuC/D RNA-binding proteins in adult rat primary sensory neurons.

HuC/D RNA-binding proteins are antigens that are specifically expressed in nerve cells. In the present study, immunocytochemistry and laser confocal microscopy were used to study the cellular distribution of HuC/D RNA-binding proteins in adult rat primary sensory neurons. Colocalization with the protein gene product 9.5 (PGP 9.5) was used to identify sensory neurons. Confocal laser imaging showed that HuC/D antigens are expressed in the cytoplasm of all rat primary sensory neurons. Most neurons also showed anti-HuC/D immuno-positivity in the nucleus. The demonstration of the ubiquitous expression of HuC/D antigens in primary sensory neurons gives us a possible explanation for the severe pathological lesions observed in dorsal root ganglia in association with the paraneoplastic Hu Syndrome.     

Physiological properties of primary sensory neurons appropriately and inappropriately innervating skin in the adult rat.

1. We have studied the physiology of sensory neurons innervating skin of the rat hindlimb, in three groups of animals: 1) normal animals; 2) animals in which the sural nerve (Sn) had regenerated to its original cutaneous target; and 3) animals in which the gastrocnemius muscle nerve (Gn) had previously been cut and cross anastomosed with the distal stump of the cut Sn so that its axons regenerated to a foreign target, skin. 2. Single-unit recordings were made from 222 afferents in normal, intact animals. They had conduction velocities of 0.5-53.1 m/s. The conduction velocity distribution had distinct peaks at approximately 37.5, 2.5, and 1.25 m/s, presumably corresponding to A alpha beta-, A delta-, and C-fiber populations. Eighty-two percent of the characterized myelinated fibers had low-threshold mechanosensitive receptive fields, whereas 16% were high threshold, and only 2% appeared to have no receptive field. The very large majority of low-threshold mechanosensitive receptive fields (87%) were rapidly adapting hair follicle afferents. 3. In animals with regenerated Sn, 308 afferents were recorded with conduction velocities of 0.4-58.8 m/s. However, the mean conduction velocity was lower than in control animals (P less than 0.05), and only one peak, at 27.5 m/s, was apparent for myelinated fibers. Eighty-six percent of myelinated fibers were low-threshold mechanosensitive afferents, 8.5% were high-threshold mechanoreceptors (HTMRs), and 5.5% appeared to have no receptive fields. Fewer low-threshold mechanoreceptors (LTMRs; compared with controls) were activated by hair movement (63 vs. 87%). Most of the remainder appeared to be field receptors (which were therefore more commonly observed here than in normal animals). 4. In animals in which the Gn had regenerated to skin, 430 afferents were recorded. These had conduction velocities ranging from 0.6 to 71.4 m/s, and again only one peak was apparent in the myelinated conduction velocity histogram, at approximately 17.5 m/s. Of the myelinated fibers, 79% had low-threshold mechanosensitive receptive fields in skin and 10% high-threshold mechanosensitive receptive fields. The remaining 11% apparently had no receptive field (cf. 5.5% in regenerated Sn). In contrast to normal or regrown sural afferents, only 58% of low-threshold gastrocnemius afferents in skin were rapidly adapting. Of the 42% slowly adapting afferents, many surprisingly responded to hair movement. Thus some gastrocnemius afferents seemed to have retained the adaptation properties characteristic of muscle afferents. Also surprisingly, given that the Gn contains fewer fibers than the Sn, receptive-field areas were not significantly different from regrown or normal sural fibers.(ABSTRACT TRUNCATED AT 400 WORDS)     

Small primary sensory neurons innervating epidermis and viscera display differential phenotype in the adult rat.

Target tissues contribute to the phenotype and function of sensory neurons. Due to lack of appropriate markers for trkA expressing sensory axons and terminals, the detailed peripheral projection of these neurons is unclear. In this study, the peripheral projections of trkA immunoreactive neurons are characterized using the combined techniques of immunohistochemistry and retrograde tracing. We found approximately 65% of all neurons projecting to the adrenal gland and kidney are trkA immunoreactive, whereas 6, 14 and 37% of neurons innervating whisker follicle, epidermis and footpad, respectively, are immunoreactive for trkA. A low proportion of trkA immunoreactive neurons innervating epidermis indicates that the majority of sensory neurons innervating epidermis are independent of trkA signalling for their normal function. We further investigated whether these epidermal projecting neurons can bind isolectin IB4. We found approximately 70% of all neurons innervating epidermis are IB4 binding neurons, but they did not express trkA. Thus, NGF sensitive neurons primarily project to viscera but not epidermis or other skin structures, whereas IB-4 positive neurons primarily project to epidermis in the adult rat.     

Proton inhibition of GABA-activated current in rat primary sensory neurons

 The modulation of the Cl^– current activated by γ-aminobutyric acid (GABA) by changes in extracellular pH in freshly isolated rat dorsal root ganglia (DRG) neurons was studied using the whole-cell patch-clamp technique. In the pH range of 5.0–9.0, increased extracellular pH enhanced, and decreased extracellular pH suppressed, current activated by 10 μM GABA in a reversible and concentration-dependent manner with an IC₅₀ of pH 7.1 in these neurons. Acidification to pH 6.5 inhibited currents activated by the GABA_A-selective agonist muscimol in all neurons tested. The antagonism of GABA-activated current by lowering the pH was equivalent at holding potentials between –80 and +40 mV and did not involve a significant alteration in reversal potential. Acidification shifted the GABA concentration/response curve to the right, significantly increasing the EC₅₀ for GABA without appreciably changing the slope or maximal value of the curve. Inhibition of the GABA-activated current by protons was not significantly different when the patch-pipette solution was buffered at pH 7.4 or pH 6.5. These results suggest that extracellular protons inhibit GABA_A receptor channels in primary sensory neurons by decreasing the apparent affinity of the receptor for GABA. This represents a novel mechanism of inhibition by protons of a neurotransmitter-gated ion channel. Proton inhibition of GABA_A receptor channels may account in part for the modulation by protons of sensory information transmission under certain pathophysiological conditions. Received: 1 July 1997 / Received after revision: 29 September 1997 / Accepted: 15 October 1997     

Purinergic synapses formed between rat sensory neurons in primary culture

Though there is some evidence to the contrary, dogma claims that primary sensory neurons in the dorsal root ganglion do not interact, that the ganglion serves as a through-station in which no signal processing occurs. Here we use patch clamp and immunocytochemistry to show that sensory neurons in primary culture can form chemical synapses on each other. The resulting neurotransmitter release is calcium dependent and uses synaptotagmin-containing vesicles. On many cells studied, the postsynaptic receptor for the neurotransmitter is a P2X receptor, an ion channel activated by extracellular ATP. This shows that sensory neurons have the machinery to form purinergic synapses on each other and that they do so when placed in short-term tissue culture.     

Bax protein-like immunoreactivity in primary sensory and hypothalamic neurons of adult rats

Bax protein-like immunoreactivity (Bax-ir) was examined in the perfusion-fixed, cryosectioned rat nervous system. In the central nervous system, hypothalamic neurons were the only neurons that exhibited Bax-ir in the cell body. Their axons traveled toward the median eminence, suggesting that the Bax-like immunoreactive (Bax-ir) hypothalamic neurons included neurosecretory ones. Bax-ir axons were observed in the solitary tract nucleus, and spinal and medullary dorsal horns. They appear to have been derived from Bax-ir primary sensory neurons in the viscerosensory nodose ganglion and somatosensory dorsal root and trigeminal ganglia. In the somatosensory ganglia, smaller cells exhibited stronger Bax-ir. Accordingly, the ir axons in the dorsal horn were most concentrated in lamina II.     

Ultrastructural localization of brain-derived neurotrophic factor in rat primary sensory neurons

In a previous study we have shown that brain-derived neurotrophic factor (BDNF) is present in a subpopulation of small- to medium-sized sensory neurons in the dorsal root ganglia (DRG) and is anterogradely transported in both the peripheral and central processes. Within the spinal cord, BDNF is localized to varicosities of sensory nerve terminals in laminae I and II of the dorsal horn. This study raised the question of whether BDNF is localized in synaptic vesicles of the afferent nerve terminals. Using immunohistochemical and immunocytochemical techniques we have now investigated the ultrastructural localization of BDNF in the spinal cord of the rat. In addition, its colocalization with the low affinity neurotrophin receptor, p75, and calcitonin gene related peptide (CGRP) was also investigated. In lamina II of the spinal cord, BDNF immunoreactivity was restricted to nerve terminals. The reaction product appeared associated with dense-cored and clear vesicles of terminals superficial laminae. Double labelling experiments at the light microscopic level showed that 55% of BDNF immunoreactive neurons in DRG are colocalized with CGRP and many nerve terminals in laminae I and II of the spinal cord contained both BDNF and CGRP immunoreactivities. The results of double labelling at the ultrastructural level showed that most BDNF-ir (immunoreactive) nerve terminals contained CGRP or the low affinity neurotrophin receptor, p75, but not vice versa. These results point to the conclusion that BDNF may be released in parallel with neurotransmitters from nerve terminals in the spinal cord from a subpopulation of nociceptive primary afferents.     

Characteristics of late Na(+) current in adult rat small sensory neurons

Na(+) currents were recorded using patch-clamp techniques from small-diameter (<25 micrometers) dorsal root ganglion neurons, cultured from adult rats (>150 g). Late Na(+) currents maintained throughout long-duration voltage-clamp steps (>/=200 ms) were of two types: a low-threshold, tetrodotoxin-sensitive (TTX-s) current that was largely blocked by 200 nM TTX, and a high-threshold, TTX-resistant (TTX-r) current. TTX-s late current was found in approximately 28% (10/36) of small-diameter neurons and was recorded only in neurons exhibiting TTX-s transient current. TTX-s transient current activation/inactivation gating overlap existed over a narrow potential range, centered between -30 and -40 mV, whereas late current operated over a wider range. The kinetics associated with de-inactivation of TTX-s late current were slow (tau approximately 37 ms at -50 mV), strongly suggesting that different subpopulations of TTX-s channel generate transient and late current. High-threshold TTX-r late current was only present in neurons generating TTX-r transient current. TTX-r late current operated over the same potential range as that for TTX-r transient current activation/inactivation gating overlap, and activation/inactivation gating overlap could be measured even after 1.5-s-duration pre-pulses.We suggest that TTX-s late sodium current results from channel openings different from those generating transient current. As in large-diameter sensory neurons, TTX-s channels generating late openings may play a key role in controlling membrane excitability. In contrast, a single population of high-threshold TTX-r channels may account for both transient and late TTX-r currents.     

Conduction velocity changes along the processes of rat primary sensory neurons

Conduction velocities of rat L4, L5 and L6 dorsal root ganglion neurons were measured in vitro, from several points on the peripheral nerve and dorsal root. Conduction velocities calculated from a single stimulation point (12–26 mm from the ganglion) proved accurate for fibres conducting up to 17 m/s in the peripheral nerve and up to 14 m/s in the dorsal root, but tended to underestimate the value for faster fibres.

C-fibre neurons of the L4 and L5 ganglia had a unimodal distribution of conduction velocities below 1.3 m/s.These were discontinuous with A-fibre conduction velocities, which also had a unimodal distribution and had no clear Aδ peak. In contrast, conduction velocities of L6 ganglion neurons showed no discontinuity between C- and A-fibres, but had a clear Aδ peak.In A-fibre neurons, dorsal root conduction velocities were on average about 14% slower than, and linearly related to, those in the peripheral nerve. However, in individual neurons the dorsal root conduction velocity could be slower or faster than that in the peripheral nerve. In C-fibre neurons dorsal root conduction velocities were almost always slower (average 28%) but not correlated with those of the peripheral nerve.

Slowing of conduction velocity along the sciatic nerve was seen in most fibres conducting at less than 2 m/s, but not in faster fibres. The slowing was substantial (up to 60%), sometimes from the Aδ to the C-fibre range, and sudden, occurring at a distance of between 15 and 29mm from the ganglion.     

Primary sensory neurons and satellite cells after peripheral axotomy in the adult rat

The timecourse of cell death in adult dorsal root ganglia after peripheral axotomy has not been fully characterised. It is not clear whether neuronal death begins within 1 week of axotomy or continues beyond 2 months after axotomy. Similarly, neither the timecourse of satellite cell death in the adult, nor the effect of nerve repair has been described. L4 and L5 dorsal root ganglia were harvested at 1-14 days, 1-6 months after sciatic nerve division in the adult rat, in accordance with the Animals (Scientific Procedures) Act 1986. In separate groups the nerve was repaired either immediately or following a 1-week delay, and the ganglia were harvested 2 weeks after the initial transection. Microwave permeabilisation and triple staining enabled combined TUNEL staining, morphological examination and neuron counting by the stereological optical dissector technique. TUNEL-positive neurons, exhibiting a range of morphologies, were seen at all timepoints (peak 25 cells/group 2 weeks after axotomy) in axotomised ganglia only. TUNEL-positive satellite cell numbers peaked 2 months after axotomy and were more numerous in axotomised than control ganglia. L4 control ganglia contained 13,983 (SD 568) neurons and L5, 16,285 (SD 1,313). Neuron loss was greater in L5 than L4 axotomised ganglia, began at 1 week (15%, P=0.045) post-axotomy, reached 35% at 2 months ( P<0.001) and was not significantly greater at 4 months or 6 months. Volume of axotomised ganglia fell to 19% of control by 6 months ( P<0.001). In animals that underwent nerve repair, both the number of TUNEL-positive neurons and neuron loss were reduced. Immediate repair was more protective than repair after a 1-week delay. Thus TUNEL positivity precedes actual neuron loss, reflecting the time taken to complete cell death and elimination. Neuronal death begins within 1 day of peripheral axotomy, the majority occurs within the first 2 months, and limited death is still occurring at 6 months. Neuronal death is modulated by peripheral nerve repair and by its timing after axotomy. Secondary satellite cell death also occurs, peaking 2 months after axotomy. These results provide a logical framework for future research into neuronal and satellite cell death within the dorsal root ganglia and provide further insight into the process of axotomy induced neuronal death.     

Axotomy-induced alterations in the electrophysiological characteristics of neurons

The electrophysiological alterations provoked by axotomy have now been studied for almost half a century, in a number of different cell types. Consequently, it is now possible to detail some common mechanisms underlying these changes and to sort out certain trends in the data. The major phenomena reviewed in this section and some possible future directions are summarized below. (1) It is now possible to advance a unified hypothesis for the effects of axotomy on the conduction velocity of myelinated fibers. The key is that axon diameter, which is directly correlated with conduction velocity, is regulated, at least in part, by neurofilament protein gene expression and transport into the axon. Thus, the largest myelinated axons, with the fastest conduction velocities, have the highest neurofilament contents, and in turn, experience greater or faster declines in neurofilament content, axon caliber, and conduction speed following nerve injury. This regulation of neurofilament gene expression also appears to be target- and/or accessory cell-dependent. In fact, Hoffman and colleagues (1988) have hypothesized that neuron interactions with specific targets (via as yet unknown target-induced signals) may either specify or permit specification of the level of neurofilament gene expression in neurons. Imposed on this primary size determinant is an influence of activity, which also underlies the differential atrophy and decrement in conduction velocity exhibited by motor and sensory fibers of comparable diameters in the same lesioned nerve. Unmyelinated axons, whose structures are not dominated by neurofilament content and metabolism, react very differently to axotomy. The structural and metabolic basis of their reaction is not known. (2) Passive membrane properties, in particular neuronal input resistance, remain relatively stable in the majority of neurons after axotomy. The major exceptions, vertebrate spinal motoneurons, lamprey dorsal interneurons, and mammalian vagal motoneurons, all show an increase in input resistance after axotomy. This change in input resistance appears to be correlated with structural or geometric simplification of dendritic trees and real or apparent changes in specific membrane resistance in one case and with a reduction in cell body size in the other two; however, changes in specific membrane resistance cannot be excluded even in the latter two cases. In the spinal motoneurons, input resistance changes may be more pronounced in those neurons with the most extensive or complex dendritic geometries (i.e. F-type motoneurons). More combined electrophysiological (ideally under voltage or patch clamp conditions) and morphological investigations of single neurons need be done to resolve these questions.(ABSTRACT TRUNCATED AT 400 WORDS)     

Injured primary sensory neurons switch phenotype for brain-derived neurotrophic factor in the rat.

Peripheral nerve injury results in plastic changes in the dorsal root ganglia and spinal cord, and is often complicated with neuropathic pain. The mechanisms underlying these changes are not known. We have now investigated the expression of brain-derived neurotrophic factor in the dorsal root ganglia with histochemical and biochemical methods following sciatic nerve lesion in the rat. The percentage of neurons immunoreactive for brain-derived neurotrophic factor in the ipsilateral dorsal root ganglia was significantly increased as early as 24 h after the nerve lesion and the increase lasted for at least two weeks. The level of brain-derived neurotrophic factor messenger RNA was also significantly increased in the ipsibut not contralateral dorsal root ganglia. Both neurons and satellite cells in the lesioned dorsal root ganglia synthesized brain-derived neurotrophic factor messenger RNA after the nerve lesion. There was a dramatic shift in size distribution of positive neurons towards large sizes seven days after sciatic nerve lesion. Morphometric analysis and retrograde tracing studies showed that no injured neurons smaller than 600 microm2 were immunoreactive for brain-derived neurotrophic factor, whereas the majority of large injured neurons were immunoreactive in the ipsilateral dorsal root ganglia seven days postlesion. The brain-derived neurotrophic factor-immunoreactive nerve terminals in the ipsilateral spinal cord were reduced in the central region of lamina II, but increased in more medial regions or deeper into laminae III/IV. These studies indicate that sciatic nerve injury results in a differential regulation of brain-derived neurotrophic factor in different subpopulations of sensory neurons in the dorsal root ganglia. Small neurons switched off their normal synthesis of brain-derived neurotrophic factor, whereas larger ones switched to a brain-derived neurotrophic factor phenotype. The phenotypic switch may have functional implications in neuronal plasticity and generation of neuropathic pain after nerve injury.     

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.     

Physiological properties of primary sensory neurons appropriately and inappropriately innervating skeletal muscle in adult rats.

1. We have studied the physiology of primary sensory neurons innervating rat hindlimb muscle in the following: 1) normal control animals; 2) animals in which the gastrocnemius nerve (Gn) had regenerated to its original muscle target; and 3) animals in which the cutaneous sural nerve (Sn) had regenerated to a foreign target, muscle. 2. Single-unit recordings were made from 115 afferents in normal, intact animals. They had conduction velocities of 0.8-67.2 m/s, which were distributed with peaks at approximately 1.25, 17.5, and 47.5 m/s. Of the myelinated fibers, 88% had low-threshold mechanosensitive receptive fields and responded to ramp-and-hold stretches of the muscle. The large majority of these fibers (85%) gave slowly adapting responses to ramp-and-hold stretches or direct muscle probing. Stretch-sensitive afferents could be divided (on the basis of their responses to active muscle contraction) into in-parallel or in-series receptors (presumed muscle spindles and Golgi tendon organs, respectively). The in-parallel receptors outnumbered the in-series receptors by approximately 3:2. The 12% of fibers that were insensitive to stretches of the muscle in the physiological range could be divided into roughly equal groups of totally insensitive fibers and high-threshold fibers, which required excessive stretching of the muscle. 3. In the animals with regrown Gn, 94 single fibers with conduction velocities ranging from 11 to 60.6 m/s were studied. The myelinated conduction velocity distribution exhibited only one peak, at approximately 37.5 m/s. Only 67% of the afferents were stretch sensitive (vs. 88% in normal animals), and only about two-thirds of these (vs. 85% in normal animals) gave slowly adapting responses to ramp-and-hold stretches or muscle probing. The incidence of in-series receptors was also increased among regenerated gastrocnemius afferents. The 33% of fibers that were stretch insensitive were mostly unresponsive to even extreme forms of mechanical stimuli. This group presumably represents afferents that failed to make appropriate endings. 4. In the animals with Sn directed to muscle, 460 single afferents were recorded. Their conduction velocities ranged from 0.7 to 67.9 m/s, and the distribution exhibited only a single peak for myelinated fibers at approximately 22.5 m/s, significantly lower than for intact or regrown Gn. Only 41% of the myelinated fibers were stretch sensitive. Nearly all of these (98%) were rapidly adapting to ramp-and-hold stretches or muscle probing, in marked contrast to the other groups. Also, unlike other groups, nearly all stretch-sensitive afferents appeared to be in-series.(ABSTRACT TRUNCATED AT 400 WORDS)     

Intracisternal capsaicin: Selective degeneration of chemosensitive primary sensory afferents in the adult rat

The present study reports that intracisternal administration of capsaicin induces the selective degeneration of chemosensitive primary sensory afferents and results in a practically complete abolition of chemical pain sensitivity in the adult rat. This treatment, however, failed to affect neurogenic inflammation in the corresponding skin areas. Accordingly, intracisternal capsaicin induces merely the degeneration of the centrally directed axons of chemosensitive primary sensory neurones (CPSNs). To indicate their particular dual function, CPSNs are proposed to be termed secreto-sensory nociceptive neurones. It is suggested that these neurones, through the release of neurogenic factor(s) at their peripheral end, may effectively modulate the afferent input related to pain sensation at the level of sensory receptors.     

Differential expression of alpha-CGRP and beta-CGRP by primary sensory neurons and enteric autonomic neurons of the rat

Expression of the calcitonin gene-related peptide, alpha-calcitonin gene-related peptide (CGRP), and the homologous beta-CGRP were compared in sensory and enteric nerves of the rat. Analysis of CGRP-like immunoreactivity by cation exchange chromatography and radioimmunoassay showed that in the dorsal root ganglia, dorsal spinal cord and in those peripheral tissues where CGRP-like immunoreactivity is primarily localized to sensory fibres, alpha-CGRP concentrations were three to six times greater than beta-CGRP concentrations. In the intestine, however, beta-CGRP concentrations were up to seven times greater than alpha-CGRP concentrations. Only beta-CGRP was detected in the intestines of capsaicin-treated rats. Northern blot and in situ hybridization to alpha-CGRP- and beta-CGRP-specific probes showed that while both alpha-CGRP and beta-CGRP messenger ribonucleic acids occurred in the dorsal root ganglia, only beta-CGRP messenger ribonucleic acid occurred in the intestine, where it was localized to enteric neurons. Receptor binding sites on membranes of rat heart and colon had approximately equal affinities for alpha-CGRP and beta-CGRP. The two peptides were equipotent in increasing the rate and force of atrial contractions but alpha-CGRP was slightly (2.6 times) more potent than beta-CGRP in relaxing colonic smooth muscle. Thus, both alpha-CGRP and beta-CGRP occur in the rat nervous system and are both biologically active. Sensory neurons and enteric neurons have been identified as populations which preferentially express alpha-CGRP and beta-CGRP, respectively.     

Mechanosensitive potassium channels in rat colon sensory neurons

Single-channel recording techniques were used to characterize mechanosensitive channels in identified (1.1'-dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine methanesulfonate labeled) colon sensory neurons dissociated from adult S1 dorsal root ganglia. Channels were found in 30% (7/23) of patches in a cell-attached configuration and in 43% (48/111) of excised inside-out patches. Channels were highly selective for K(+), had a slope conductance of 54 pS in symmetrical solutions, and were blocked by tetraethylammonium, amiloride, and benzamil. Channels were also seen under Ca(2+)-free conditions. Gadolinium (Gd(3+)), a known blocker of mechanosensitive ion channels, did not block channel activity. Tetrodotoxin and 4-aminopyridine were also ineffective. The cytoskeletal disrupters colchicine and cytochalasin D reduced the percentage of patches containing mechanosensitive channels. These results indicate that rat colon sensory neurons contain K(+)-selective mechanosensitive channels that may modulate the membrane excitability induced by colonic distension.