Functional and morphological consequences of kainic acid microinjections into a pontine respiratory area of the cat

The functional and morphological effects of kainic acid microinjections have been investigated in the nucleus parabrachialis medialis, a pontine respiration-related structure. Individual sensitivity of respiratory-modulated neurones located in the nucleus parabrachialis medialis tol-glutamate (an excitatory aminoacid analog to kainic acid) was demonstrated using microiontophoretic applications and extracellular recordings. After bivagotomy a small dose of kainic acid (0.5 μg in 0.5 μl) injected into one nucleus parabrachialis medialis induced irreversible apneustic breathing similar to that observed after electrocoagulation of the nucleus parabrachialis medialis. When the vagi were intact the same microinjection led to a complete cessation of spontaneous breathing of barbiturate-anaesthetized cats. Such irreversible effects were not reproduced in control experiments with comparable injections ofl-glutamate or Ringer's solution in the nucleus parabrachialis medialis or with kainic acid injections outside the nucleus parabrachialis medialis. In another experimental series, after bivagotomy, large doses of kainic acid (4–6 μg in 8–12 microinjections on each side) were applied in a transverse plane caudal to the nucleus parabrachialis medialis; no apparent respiratory change was observed while electrocoagulation performed at the same level caused apneusis. The anatomical data obtained under light and electron microscopy after kainic acid microinjections have shown a rapid action upon various components of nervous tissue. There is a severe neuronal loss at the level of the injection site. Two hours after injection, somata exhibit the first signs of degeneration, but also axons and glia begin to react. At 20 h survival degeneration in all components continues to increase.The functional and morphological sequelae of kainic acid are discussed. Its usefulness as a neuropharmacological tool is confirmed but limits to its specificity have been found.     

Wheel-running activity after kainic acid injection into lateral hypothalamus of rats

It has not been resolved whether the permanent decrease in wheel-running activity observed after the placement of bilateral electrolytic lesions in the ventrolateral hypothalamus of rats is due to local neuronal destruction or to disruption of fibers of passage within the lateral hypothalamus. To further explore this question, the changes in wheel-running activity following injections of a kainic acid (KA) solution into the ventrolateral hypothalamus of rats were studied. As was found with electrolytic lesions, KA induced lesions in the ventrolateral hypothalamus resulted in a permanent decrease in wheel-running activity. The uptake of 3H-dopamine (3H-DA) into crude synaptosomal preparations of striatal tissue was used as an index of the amount of damage done to fibers of passage by KA. 3H-DA uptake by striatal tissue from rats injected with KA did not significantly differ from that of control rats. These data support the hypothesis that the decrease in wheel-running activity following injection of KA into the ventrolateral hypothalamus is the result of damage to intrinsic neurons.     

Capsaicin infused into the PAG affects rat tail flick responses to noxious heat and alters neuronal firing in the RVM

It is well established that the vanilloid receptor, VR1, is an important peripheral mediator of nociception. VR1 receptors are also located in several brain regions, yet it is uncertain whether these supraspinal VR1 receptors have any influence on the nociceptive system. To investigate a possible nociceptive role for supraspinal VR1 receptors, capsaicin (10 nmol in 0.4 microl) was microinjected into either the dorsal (dPAG) or ventral (vPAG) regions of the periaqueductal gray. Capsaicin-related effects on tail flick latency (immersion in 52 degrees C water) and on neuronal activity (on-, off-, and neutral cells) in the rostral ventromedial medulla (RVM) were measured in lightly anesthetized rats. Administration of capsaicin into the dPAG but not the vPAG caused an initial hyperalgesic response followed later by analgesia (125 +/- 20.96 min postinjection). The tail flick-related burst in on-cell activity was triggered earlier in the hyperalgesic phase and was delayed or absent during the analgesic phase. Spontaneous activity of on-cells increased at the onset of the hyperalgesic phase and decreased before and during the analgesic phase. The tail flick-related pause in off-cell activity as well as spontaneous firing for these cells was unchanged in the hyperalgesic phase. During the analgesic phase, off-cells no longer paused during noxious stimulation and had increased levels of spontaneous activity. Neutral cell firing was unaffected in either phase. Pretreatment with the VR1 receptor antagonist, capsazepine (10 nmol in 0.4 microl), into the dPAG blocked the capsaicin-induced hyperalgesia as well as the corresponding changes in on- and off-cell activity. VR1 receptor immunostaining was observed in the dPAG of untreated rats. Microinjection of capsaicin likely sensitized and then desensitized dPAG neurons affecting nocifensive reflexes and RVM neuronal activity. These results suggest that supraspinal VR1 receptors in the dPAG contribute to descending modulation of nociception.     

Different sensitivity of hippocampal neurons to kainic acid in the rabbit

The toxic effect of kainic acid, a cyclic analogue of glutamic acid, was investigated in the rabbit hippocampus. Using histological methods it was found that intrahippocampal injection of one or two doses of 5 nm of kainic acid produced cell losses in different cell layers. The cells with the highest sensitivity to kainic acid were the cornu ammonis pyramidal cells, whereas those with the lowest sensitivity were the dentate granular cells.These findings suggest that the sensitivity to kainic acid is different in discrete cell areas of the hippocampus.     

Effects of LH kainic acid infusions on ingestion and autonomic activity

The effects of lateral hypothalamic (LH) infusions of kainic acid (KA) were determined on ingestive behavior, body weight, and motor and autonomic activity. In Experiment 1 male hooded rats received bilateral LH infusions of isotonic saline in volumes of 0.5 or 1.0 μl, 3.0 μg KA in volumes of 0.5 or 1.0 μl, or 6.0 μg of KA in a volume of 1.0 μl. All animals receiving 6.0 μg/1.0 μl of KA died. The 3.0 μg/0.5 μl dose resulted in transient decreases in food and water consumption and body weight. Animals receiving this dose no longer drank in response to 2 cc/kg 15% NaCl injections, exhibited attenuated drinking in response to 24 hr water and food deprivation, exhibited a transient decrease in eating following food deprivation and decreased eating following 750 mg/kg injections of 2-deoxy-D-glucose. Minimal effects on these measures were observed following the 3.0 μg/1.0 μl dose. In Experiment 2 rats received unilateral infusions of KA and the effects on motor and autonomic activity and ingestive behavior and body weight were compared to unilateral saline infused animals and animals with radiofrequency lesions. Only transient decreases in food consumption lasting 1–2 days were observed for both unilateral KA LH infused and lesioned animals. In KA infused rats contralateral exophthalamus, rapid shallow breathing, bilateral mydriasis, no contralateral pupillary constriction response, excessive salivation, body tremors, seizures, convulsions, teeth chattering, contralateral tail suspension induced spinning and turning, and elevated body temperature were observed for up to 6 hr following the infusion. Results are discussed in terms of lateral ventral diencephalic neurons involved in the more permanent deficits associated with bilateral LH damage.     

Maturation of kainic acid seizure-brain damage syndrome in the rat. III. Postnatal development of kainic acid binding sites in the limbic system

The progressive appearance of [3H]kainic acid binding sites with age has been studied in membrane suspensions prepared from various regions of the rat limbic system, and by autoradiography. Binding sites with fast dissociation rate appeared earlier than binding sites with slow dissociation rate. Scatchard analysis demonstrated apparent receptor heterogeneity for both subclasses. High affinity components were detected in the hippocampus as early as 10 days after birth, but in the amygdala + piriform lobe were found only towards the end of the third week, when animals also respond to parenteral kainic acid, for the first time, with limbic seizures accompanied by metabolic activation of the amygdala. Slice autoradiography revealed distinct labelling of the hippocampal CA3 region by postnatal day 10. A comparison with the ontogenesis of the kainic acid-induced seizure-brain damage syndrome suggests a role of high affinity receptors as mediators of metabolic nerve cell activation by kainic acid. However, this receptor interaction per se does not result in neuronal damage to the vulnerable region of the Ammon's horn, which will only occur at an age when also the amygdala is activated by the neurotoxin.     

Differential effect of kainic acid on somatostatin, GABAergic and cholinergic neurons in the rat striatum

Kainic acid was injected into the rat striatum and its effects on presumptive striatal cholinergic, GABAergic and somatostatin-containing neurons were examined with three histochemical staining methods. Presumptive cholinergic and GABAergic neurons are damaged to a similar extent, but somatostatin neurons are more sensitive to the neurotoxic effect of kainic acid and are more severely affected.     

Effects of neuronal activity on kainic acid neurotoxicity in the ventral cochlear nucleus

Kainic acid was injected into the brain stem of adult guinea pigs, and the animals were either placed in a sound reducing-chamber or stimulated with 90 dB noise. The pattern and rate of kainic acid-induced degeneration in the anteroventral cochlear nucleus (AVCN) of sound-deprived animals was similar to that in animals exposed to ambient noise [2]. The amount of degeneration was greatly increased in animals stimulated with 90 dB noise. Therefore, although decreased activity in primary auditory fibers does not protect neurons in the AVCN from kainate-induced neurotoxicity, increased auditory stimulation augments the effects of kainic acid in the cochlear nucleus.     

红藻氨酸对大鼠海马锥体细胞钙电流的影响

目的：应用膜片钳技术,观察红藻氨酸（KA）对大鼠海马锥体细胞ca2＋电流的影响,以研究癫痫的发病机制。方法：采用酶加机械分离法制备出生10～12d的大鼠海马锥体神经元标本,用全细胞膜片钳技术测定其生理学特性及观察KA对ca2＋电流的影响。结果：分离出的海马锥体细胞形态正常,有较长突起;用膜片钳技术证实,其保存了主要的离子通道活性。KA20μmol／L和100μmol／L的浓度均可使海马锥体细胞ca2＋电流峰值增大（n=8,P〈0．01）。结论：①大鼠海马锥体神经细胞具有明显的突起,细胞膜表面光洁、晕光好,适用于膜片钳实验研究;②KA使ca2＋内流增加,引起“Ca”超载”导致细胞毒性等一系列反应;③KA通过激活α-氨基羟甲基恶唑丙酸（AMPA）受体,诱发快速的兴奋性突触后电位（EPSP）,参与兴奋性突触传递。AMPA受体的激活可能是癫痫的发病机制之一。     

Integrated excitatory response to microinjections of L-glutamic acid into the reticular system of the toad

Microinjections of L-glutamic acid into the mesencephalic tegmentum and diencephalon of anesthetized toads evoked an excitatory response consisting of electrocephalographic and electromyographic activation, and a rise in arterial blood pressure. Electromyographic responses were reduced or absent in diencephalically injected toads. Except for this case, all other components of the response always appeared together and presented the same time course. L-glutamic acid was able to activate a group of neurons at different levels of the reticular system involved in an integrated response of ergotropic nature.     

Effects of acetylsalicylic acid and morphine on neurons of the rostral ventromedial medulla in rat

Morphine exerts its analgesic effect via the endogenous pain control system consisting of the periaqueductal grey (PAG) and the rostral ventromedial medulla (RVM). Acetylsalicylic acid (ASA) may also act via this system, but so far this has only been demonstrated for the inhibitory effect on the tail-flick reflex with extremely high doses (200-300 mg/kg). Both drugs show synergistic effects on PAG neurons in vitro. It is unclear whether this mechanism accounts for the well-known analgesic synergism of these drugs in vivo. Thus, the effects of ASA (30 mg/kg) and morphine on off- and on-cells in the RVM and the jaw-opening reflex (JOR) were investigated in anesthetized rats. Under morphine, off-cell activity increased (+34%), on-cell activity decreased (-98%) and the reflex was suppressed (-53%). ASA increased off-cell activity (+20%) and decreased the activity of on-cells (-52%). After preceding ASA administration, the effects of morphine on off- and on-cells and on the reflex did not alter statistically. The experiments document the modulatory effect of a clinically relevant dose of ASA on RVM cells. This effect resembles that of morphine. The results do not support the hypothesis of a mediation of the analgesic synergism of morphine and ASA by the PAG-RVM-network in vivo.     

Tolerance to the antinociceptive effect of morphine microinjections into the ventral but not lateral-dorsal periaqueductal gray of the rat.

Tolerance to the antinociceptive effect of morphine is mediated at least in part by morphine's action within the periaqueductal gray (PAG). The objective of the present study was to determine whether both ventral and lateral-dorsal PAG regions contribute to the development of tolerance. It was found that the antinociceptive efficacy of microinjecting morphine (5 microg/0.4 microl) into the ventral but not the lateral-dorsal PAG diminished with successive injections. Control experiments indicated that this decrease was caused by tolerance to morphine and was not a result of cell death caused by repeated microinjections or habituation from repeated behavioral testing. The finding of greater susceptibility of the ventral compared with the lateral-dorsal PAG to the development of tolerance adds to a growing literature distinguishing antinociception from these two regions.     

Comparative toxicity of kainic acid and other acidic amino acids toward rat hippocampal neurons

Acidic amino acids were tested by histological criteria for their ability to destroy rat hippocampal neurons. When injected directly into the hippocampal formation, kainic acid preferentially destroyed CA3-CA4 pyramidal cells and was least toxic toward dentate granule cells and pyramidal cells of the h₂ area. All other acidic amino acids tested destroyed hippocampal neurons non-selectively, the vulnerability of these neurons depending simply on their proximity to the injection site. At sufficient doses, kainic acid destroyed neurons in distant limbic regions, but the other acidic amino acids did not. All amino acid excitants tested were neurotoxic, the order of potency being:kainate>ibotenate>N-methyl-dl-aspartate>dihydrokainate>dl-homocysteate>l-cysteate>l-aspartate>l-glutamate. Most excitatory amino acid antagonists were only very weakly neurotoxic, but the phosphonate analogues of glutamate and aspartate were about as potent as the strong excitants,dl-homocysteate andl-cysteate. Intraventricular injections similarly revealed a distinction between selective hippocampal lesions made by kainic acid and non-selective hippocampal lesions made by high doses of other acidic amino acids.These results generally support previous correlations between neuro-excitatory and neurotoxic effects of acidic amino acids. However, the considerable neurotoxic potency of the phosphonate antagonists is difficult to reconcile with the conventional view that the ability of acidic amino acids to destroy neurons is determined solely by their ability to depolarize them. Several mechanisms are discussed to account for the different patterns of neuronal cell death observed after injection of kainic acid and other acidic amino acids. Finally,N-methyl-dl-aspartate is suggested as a useful agent for making localized axonsparing lesions.     

Effect of intra-cisternal application of kainic acid on the spinal cord and locomotor activity in rats

Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease of idiopathic etiology. Glutamate excitotoxicity is one of the proposed hypotheses causing progressive death of motor neurons. We aimed to develop an experimental animal model of this disease to enhance the knowledge of pathophysiological mechanism of ALS. Male Wistar rats were infused with Kainic acid (KA) intra-cisternally for 5 days at the dosage of 50 fmol/day and 150 fmol/day. Locomotor activity, sensory function and histological changes in cervical and lumbar sections of spinal cord were evaluated. Glial Fibrillary Acidic Protein (GFAP) and Neurofilament Protein (NFP) were used as immunohistochemical marker for reactive astrogliosis and neuronal damage respectively. Specific Superoxide Dismutase (SOD) activity of spinal cord was estimated. The locomotor activity in the parameter of observed mean action time remained reduced on 14^th day after administration of KA. Spinal motor neurons under Nissl stain showed pyknosis of nucleus and vacuolation of neuropil. GFAP expression increased significantly in the lumbar section of the spinal cord with high dose of KA treatment (p<0.05). NFP was expressed in axonal fibres around the neurons in KA-treated rats. A significant increase in specific SOD activity in both cervical and lumbar sections of the spinal cord was found with low dose of KA treatment (p<0.05). This study concludes that spinal cord damage with some features similar to ALS can be produced by low dose intra-cisternal administration of KA.     

Involvement of local orphanin FQ in the development of analgesic tolerance induced by morphine microinjections into the dorsal raphe nucleus of rats

It is well known that dorsal raphe nucleus (DRN) is one of the key structures for the development of opioid analgesia and tolerance. An increased activity of ‘antiopioids’ like orphanin-FQ (OFQ) has been proposed as a possible mechanism for opioid tolerance. The present study evaluates the role of DRN-located OFQ in the opioid analgesic tolerance induced by repeated microinjections of morphine (MOR) into DRN. Male rats were implanted with chronic guide cannulae aimed at the DRN. Microinjection of MOR (0.5 μg in 0.5 μl) into DRN caused antinociception as quantified with the tail flick and the hot plate tests. When MOR microinjection was repeated twice daily, the antinociceptive effect disappeared within 2 days (tolerance). However, if each MOR microinjection was preceded (within 15 min) by a microinjection of the OFQ receptor antagonist nocistatin (NST) (1 ng in 0.5 μl) into the same DRN site, the microinjections of MOR always produced antinociception and did not induce tolerance. If NST microinjections were suspended, subsequent MOR microinjections induced tolerance. In MOR-tolerant rats, a single NST microinjection into the same DRN site was enough to restore the antinociceptive effect of MOR. On the other hand, if OFQ (1 ng in 0.5 μl) was microinjected into DRN, then MOR microinjection administered 15 min later into the same DRN site did not elicit antinociception. Finally, opioid tolerance induced by repeated systemic MOR injections (5 mg/kg, ip) was reversed by a single microinjection of NST into DRN. This emphasizes the central importance of DRN-located OFQ in the MOR analgesic tolerance.     

The actions of cyclopentane analogues of glutamic acid at binding sites for kainic and glutamic acids

Summary The actions of the four isomers of 1-amino-1, 3-cyclopentane dicarboxylate (ACPD), a conformationally restricted analogue of glutamate, have been examined for their ability to displace radiolabelled kainate and glutamate from their binding sites on membranes prepared from rat brain. All four of the isomers reduced specific kainate binding, and all enhanced that of glutamate although one (D-(-)-cis-(1R,3R)-ACPD) was significantly more active in this respect than were the other three. The results are discussed in terms of the pharmacological effects of the isomers of ACPD on single neurones and the possible structural requirements of the NMDA and kainate receptors.