Supersensitivity to substance P after dorsal root section.

Two to three weeks after unilateral dorsal root section, specific [³H] substance P binding in synaptic membranes from rabbit spinal cord increased significantly compared with that in intact side, while non-specific [³H] substance P binding was unchanged. Scatchard analysis indicates that the development of such supersensitivity is due to an increase in a number of receptor sites, not to an increase in a receptor affinity. These results strongly support the notion that substance P exerts a transmitter role for primary afferent fibers.     

电生理学特性得到确定的大鼠脊髓背根神经节细胞内谷氨酸和P物质的共存

在离体灌流带脊髓和坐骨神经经的标本上,对脊髓背根神经节（ＤＲＧ）细胞的电生理学特性、对Ｐ物质（ＳＰ）受体激动剂的反应及谷氨酸（Ｇｌｕ）／ＳＰ共存的特点进行了研究。（１）对１３５个细胞进行了细胞内记录,并依纤维传导速度将其分为Ａα／β（〉１２ｍ／ｓ）和Ｃ（〈１．３ｍ／ｓ）两大类,它们的动作电位的快速后超极化（ｆＡＨＰ）有明显区别,Ｃ类的ｆＡＨＰ幅度小、时程长,Ａα／β类的ｆＡＨＰ幅度大、时程短;（２     

Supersensitivity to a substance P analogue following dorsal root section

Fourteen - twenty-one days after unilateral dorsal root section, interneurones in the deafferented side of the rat spinal cord were more sensitive to iontophoretic application of Eledoisin Related Peptide (ERP - a Substance P analogue). The cells were not more sensitive to all excitatory neurotransmitters as responses to 5-HT were similar on both sides of the cord. The increased responsiveness to ERP was not simply a result of a change in the spontaneous activity of the cells. ERP responsive cells were encountered at similar depths on both sides of the cord and therefore are likely to represent similar populations. It was concluded that supersensitivity, probably of postsynaptic origin, had occurred. These observations provide further support for the postulated role for substance P as a neurotransmitter in primary afferent neurones.     

Ventral root afferent and dorsal root efferent fibres in dog and human lower sacral nerve roots.

Two pairs of wire electrodes were used to record single afferent action potentials from ventral roots and single efferent action potentials from dorsal roots of dogs and humans. A human lower sacral ventral root contained about 20 to 30% afferents among fibres with a diameter larger than 5 microns; a comparable ventral root of a dog contained about 1% afferents. Human S3, S4 and S5 dorsal roots contained 3, 18, and 20 to 30% efferent fibres respectively; a comparable dorsal root of the dog contained less than 1% efferent fibres. Primary and secondary muscle spindle afferents, Golgi tendon organ afferents, and afferents from the mechanoreceptors of the urinary bladder and anal canal mucosa were activated in a dog ventral root by pulling bladder and anal catheters. Their peak group conduction velocities were 82, 57, 71 and 18 m/s at 34 degrees C respectively. The dog afferents conducted more than 30% faster than did comparable human nerve fibres. By strongly pulling the bladder catheter, the static human dorsal root gamma 21-motoneurons increased their activity for about 7 s which in turn strongly increased the dorsal root spindle afferent activity for more than 10 min; the human static intrafusal gamma-motoneurons seemed to show cumulative properties.     

Immunohistochemical analysis of substance P containing nerve fibres and their contacts with mast cells in the diabetic rat's tongue

Sensory neuropathy is common symptom of the diabetes mellitus and the prevalence of oral lesions is higher in diabetic patients. The distribution of substance P was studied immunohistochemically in streptozotocin induced diabetic rat's tongue. The morphological association of sensory nerves (substance P immunoreactive) with mast cells (nerve fibre-mast cell contact) was monitored. The substance P nerve fibre mast cell contacts were very scanty in control tongue. The number of substance P nerve terminals and mast cells was significantly increased (p < 0.05) in diabetes mellitus after 4 weeks of the treatment compared with the control tongue. The number of mast cell nerve contacts was even more significantly increased (p < 0.001) in diabetes. The distance between nerve fibres and mast cells was about 1 mm and very often less than 200 nm. In some instances, the mast cells were degranulated in the vicinity to nerve fibres. Increased number of mast cell nerve contacts in neurogenic inflammation might cause vasoconstriction and lesions of the oral mucosa, so some disorders such lichen planus, leukoplakia and cancer might frequently develop in diabetes mellitus.     

Activation of the neurokinin-1 receptor by substance P triggers the release of substance P from cultured adult rat dorsal root ganglion neurons

Hypoxia and ischemia occur in the spinal cord when blood vessels of the spinal cord are compressed under pathological conditions such as spinal stenosis, tumors, and traumatic spinal injury. Here by using spinal cord slice preparations and patch-clamp recordings we investigated the influence of an ischemia-simulating medium on dorsal horn neurons in deep lamina, a region that plays a significant role in sensory hypersensitivity and pathological pain. We found that the ischemia-simulating medium induced large inward currents in dorsal horn neurons recorded. The onset of the ischemia-induced inward currents was age-dependent, being onset earlier in older animals. Increases of sensory input by the stimulation of afferent fibers with electrical impulses or by capsaicin significantly speeded up the onset of the ischemia-induced inward currents. The ischemia-induced inward currents were abolished by the glutamate receptor antagonists CNQX (20 μM) and APV (50 μM). The ischemia-induced inward currents were also substantially inhibited by the glutamate transporter inhibitor TBOA (100 μM). Our results suggest that ischemia caused reversal operation of glutamate transporters, leading to the release of glutamate via glutamate transporters and the subsequent activation of glutamate receptors in the spinal dorsal horn neurons.     

Effects of substance P analogues on spinal dorsal horn neurons

The effects of iontophoretically applied (D-Pro2, D-Phe7, D-Trp9)-SP and (D-Pro2, D-Trp7,9)-SP on the spontaneous and evoked activity of functionally identified cat spinal dorsal horn neurons have been investigated in vivo by means of extracellular single unit recording technique. In addition, the rat spinal cord slice preparation has been used to study the actions of (D-Pro2, D-Trp7,9)-SP and (D-Arg1, D-Pro2, D-Trp7,9, Leu11)-SP on the resting membrane potential of dorsal horn neurons and also on their responses to dorsal root stimulation and exogenous SP application. We have observed that both (D-Pro2, D-Phe7, D-Trp9)-SP and (D-Pro2, D-Trp7,9)-SP produced an excitation of about 15% of all neurons tested and had a weak antagonistic effect against SP in the cat spinal cord. (D-Pro2, D-Trp7,9)-SP suppressed the SP-induced excitation in 63% of examined cells. In addition, depression of the glutamate-induced excitation and spontaneous activity was evident in 10% and 19% of the cat dorsal horn neurons tested, respectively. In the spinal cord slice preparation (D-Arg1, D-Pro2, D-Trp7,9, Leu11)-SP proved to be a more potent antagonist of the SP-induced depolarization and the dorsal root-elicited slow depolarization, if compared with (D-Pro2, D-Trp7,9)-SP.     

Immunohistochemical characterisation of dorsal root ganglia neurons supplying the porcine mammary gland

The present study investigated the chemical coding of mammary gland-projecting dorsal root ganglia (DRG) neurons using double-labelling immunohistochemistry. Earlier investigations revealed the presence of Fast blue - positive (FB+) neurons in Th9-Th12 DRG after injection of the tracer into the second, right thoracic mamma. Neurons projecting to the last right abdominal mamma were found in L1-L3 DRG. In the present study, the cryostat sections from these ganglia were stained for calcitonin gene-related peptide (CGRP), substance P (SP), nitric oxide synthase (NOS), galanin (GAL) and pituitary adenylate cyclase activating polypeptide (PACAP). Immunohistochemistry revealed that the vast majority of FB+ mammary gland-projecting neurons contained immunoreactivity to CGRP (68.87±0.7%), SP (63.4±0.9%), NOS (32.47±0.9%), GAL (16.28±0.8%) and less numerous nerve cells stained for PACAP (5.87±0.5%). The present results largely correspond with findings dealing with immunohistochemical characterization of nerve fibres supplying porcine mammary gland structures described earlier.     

Immunohistochemical studies on GABAergic neurons in the rat locus coeruleus, with special reference to their relationship to astrocytes

Purified antisera against GABA were prepared. A few small GABAergic neurons in the rat locus coeruleus were immunohistochemically demonstrated by both the unlabeled peroxidase-antiperoxidase method and the avidin-biotin peroxidase complex method using affinity-purified GABA antibodies. The glial fibrillary acidic protein immunoreactivity in this nucleus was localized by the latter method in the astrocytal framework encircling medium-sized and small neurons as well as in straight processes. Astrocytes may play a role as energy donors to these neurons.     

Selective association of nerve fibres immunoreactive for substance P or bombesin with putative cholinergic neurons of the male rat major pelvic ganglion

The male rat major pelvic ganglion contains both sympathetic and parasympathetic neurons that supply the lower urinary and digestive tracts, and the reproductive organs. The aim of this study was to describe the distribution and identify potential targets of sensory and intestinofugal axons in this ganglion. Two putative markers of these projections were chosen, substance P for primary sensory axons and bombesin for myenteric intestinofugal projections. Varicose substance P-immunoreactive axons were associated only with non-noradrenergic (putative cholinergic) somata, and most commonly with those that contained vasoactive intestinal peptide. Immunoreactivity for substance P was also present in a small group of non-noradrenergic somata, many of which were immunoreactive for enkephalins, neuropeptide Y or vasoactive intestinal peptide. Bombesin immunoreactivity was found only in preterminal and terminal (varicose) axons, the latter of which were exclusively associated with non-noradrenergic somata that contain neuropeptide Y-immunoreactivity. Some varicose axons containing either substance P-or bombesin-immunoreactivity were intermingled with clumps of small, intensely fluorescent cells. These studies indicate that substance P-and bombesin-immunoreactive axons are likely to connect with numerically small, but discrete, populations of pelvic neurons.     

The ontogenesis of monoaminergic nerve fibres in the hypophysis of Rana temporaria with special reference to the pars distalis

Summary In Rana temporaria tadpoles, fluorescent fibres appear in the prospective eminentia mediana, the pars intermedia and the pars distalis at Gosner's stage 25. During prometamorphosis the amount of fluorescent material increases around the developing primary capillary plexus in the eminentia mediana. In the pars intermedia the fibres form a dense fluorescent network but in the pars distalis the fibres are few and delicate.At stages 42–43, the onset of climax, the pars distalis fibres disappear. The possible functional significance of the pars distalis fibres is discussed.The background adaptation ability appears at stages 28–29, while the fluorescent pars intermedia innervation is observable at stage 25.     

Vascular effects of substance P change synaptic responsiveness of cat dorsal horn neurons

The effects of systemically administered substance P were examined on the responses induced by noxious (radiant heat) and non-noxious (air jets) peripheral stimuli in dorsal horn neurons of the feline spinal cord. Substance P produced a significant fall in arterial blood pressure and selectively enhanced responses to noxious heat stimulation. Other vasoactive substances administered systemically caused either selective increases or decreases in noxious heat induced responses which coincided with decreases or increases in systemic arterial blood pressure respectively. It was concluded that the selective neuronal effects of substance P were secondary to changes in vascular perfusion in the area stimulated by the radiant heat source.     

Glutamate-immunoreactivity in the trigeminal and dorsal root ganglia, and intraspinal neurons and fibres in the dorsal horn of the rat

Summary Putative aspartergic and glutamatergic sensory neurons in the rat were identified by autoradiography and immunocytochemistry respectively. Approximately 3% of large L₄ dorsal root ganglion neurons (diameter 18–52 m) accumulated radiolabelled aspartate, whereas all satellite glia had high affinity for the amino acid. Glutamate-immunofluorescent (Glu-FITC) dorsal root ganglia neurons comprised 38.3% at S₁, 35.6% at L₂ 33.9% at C₅ and 28.8% at T₆. Numbers of immunoreactive neurons were higher with the more sensitive peroxidase-anti-peroxidase (Glu-PAP) method; and the cell counts totalled 42% (S₁), 41.2% (L₄), 35% (C₅) and 34.6% (T₆). The trigeminal ganglion (TG) contained 24% Glu-FITC and 32.3% Glu-PAP positive cells. The majority of glutamate-immunoreactive sensory neurons were small, ranging from 10–35 m with median diameters of 17.5m (C₅), 21m (S₁), 24.2m (TG) and 28.5 m (L₂). It is evident therefore, that a subgroup of class B cells are glutamatergic. Glutamate immunoreactivity in the spinal cord was similar in all segments and was localized in the superficial lamina and substantia gelatinosa of the dorsal horn. Stained interneurons were located among the immunoreactive fibres. The dorsolateral funiculus contained dense plexus of immunoreactive fibres which increased in prominence after intraperitoneal injection of L-glutamate, but penetration of exogenous glutamate into the grey matter was limited. Instead, the meninges and basal layers of the spinal blood vessels were intensely immunoreactive. The studies describe the subtypes of acidic amino acidergic neurons and relates the immunohistochemistry to a functional subclass.     

High levels of mRNA coding for substance P, somatostatin and alpha-tubulin are expressed by rat and rabbit dorsal root ganglia neurons

Oligonucleotide probes complementary to alpha-tubulin, preprotachykinin A (PPT A), preprosomatostatin (PPSOM), and preproarginine-vasopressin (PPAVP) mRNA were hybridized to sections of rat and rabbit brain and dorsal root ganglia (DRG) at all spinal levels. Approximately 100% of the DRG neurons in the rat and rabbit express alpha-tubulin mRNA, 20-30% express PPT A mRNA and 5-17% express PPSOM mRNA. Whereas neurons which express PPSOM mRNA are of relative uniform size, the neurons which express PPT A mRNA segregate into two broad groups. One group is composed of smaller neurons (200-2,000 microns 2) which contain an extremely dense concentration of PPT A mRNA. The second group is composed of larger neurons (2,000-3,500 microns 2) which contain a moderate concentration of PPT A mRNA. PPAVP mRNA is present in very high concentrations in the paraventricular and supraoptic nucleus of the rat hypothalamus but is not detected in any DRG neurons. In both the rat and the rabbit the density of PPT A and PPSOM mRNA is high in individual DRG neurons in comparison to PPT A and PPSOM mRNA levels contained in most forebrain neurons. These results suggest that although the level of neuropeptide present in DRG neurons is relatively low in comparison to other brain areas, the rate of sensory neuropeptide synthesis and turnover, as reflected by mRNA content, is extremely high.     

Neutrophic influence of denervated sciatic nerve of on adult dorsal root ganglion neurons

Isolated adult frog dorsal root ganglion neurons survive in vitro in a defined medium for more than 4 weeks and extend processes. When co-cultured with a 1-mm piece of peripheral nerve the average tottal process lenght per neuron was 10 times longer than that of control neurons by 8 days, and the processes had a significantly different morphology from that of control neurons. This influence on process length increased with increasing time of nerve denervation length increased with increasing time of nerve denervation prior to co-culturing. These results suggest the release of a neurotrophic factor/s from the cells of the peripheral nerve. The neurotropic influence was completely blocked by antibodies against mouse nerve growth factor (NGF). Although NGF increased the average process length by twofold over control neurons, its influence never reached that of the nerve-released factor, and the NGF-induced processes had a distinctly different morphology. The frog nerve-released factor promoted process outgrowth from E11 chick sympathetic ganglia, although the process number, length, and their fasciculation differed greatly from those induced by NGF. These results suggest that the nerve-released factor/s are immunologically and functionally related to NGF but have not estabished whether a single factor or an aggregate of several secreted molecules are responsible. This article presents a new preparation in which the varied influences of different neurotrophic factors can be studied in great detail on large populations of isolated adult vertebrte neurons and sets the stage for the characterization and isolation of the frog peripheral nerve neurotrophic factor, as well as examining the influence of this facor on neuronal morphology and its ability to direct process outgrowth. 1994 John Wiley & Sons, Inc.     

Influence of factors released from sciatic nerve on adult dorsal root ganglion neurons

Previous experiments have shown that medium conditioned (CM) by denervated peripheral nerve contains a process outgrowth promoting factor (s) for cultured adult frog dorsal root ganglion (DRG) neurons. The present experiments further characterize the influences of these factors on DRG neurons. The growth factors increases average process length by threefold, restricts the number of processes extended from four to two while simultaneously altering the morphology of those processes. Neurons with preexisting processes respond to the factors by significantly increasing the length of 35% of these processes. Only the newly elongated portions of preexisting processes have morphology typical of factor-induced processes, while the previously extended portions retain their original morphology. The number of processes of these neurons remains unchanged. Although composed of two population according to size, neurons in both populations are similarly influenced, suggesting that the factors influence neurons of all sensory modalities. To look at other possible influences of the nerve-released factors, a novel simple culture system has been developed in which concentration gradients of these factors can be established and maintained. The front of the outgrowth-promoting influence in these cultures could be followed over time (up to 9 days) as it affected the process length and morphology of neurons at increasing distances (up to 8 mm) from the source of the factors. The trophic factors may play important roles during regeneration in vivo by influencing the cytoskeletal organization in the cell body and growth cones to bring about a stabilization and consolidation of growth cone membrane of only a limited number of processes resulting in increasing the rate of process elongation. The factors may also serve to direct process outgrowth, which can be examined using the new culture system. 1994 John Wiley & Sons, Inc.