The Role of the Peripheral Chemokine, CCL3, in Hyperalgesia following Peripheral Nerve Injury in the Rat / 대한통증학회지

BACKGROUND: Upregulation of one type of the pro-inflammatory chemokine (CCL2) and its receptor (CCR2) following peripheral nerve injury contributes to the induction of neuropathic pain. Here, we examined whether another type of chemokine (CCL3) is involved in neuropathic pain. METHODS: We measured changes in mechanical and thermal sensitivity in the hind paws of naive rats or rats with an L5 spinal nerve ligation (SNL) after intra-plantar injection of CCL3 or met-RANTES, an antagonist of the CCL3 receptor, CCR1. We also measured CCL3 levels in the sciatic nerve and the hind paw skin as well as CCR1 expression in dorsal root ganglion (DRG) cells from the lumbar spinal segments. RESULTS: Intra-plantar injection of CCL3 into the hind paw of naive rats mimicked L5 SNL-produced hyperalgesia. Intra-plantar injection of met-RANTES into the hind paw of rats with L5 SNL attenuated hyperalgesia. L5 SNL increased CCL3 levels in the sciatic nerve and the hind paw skin on the affected side. The number of CCR1-positive DRG cells in the lumbar segments was not changed following L5 SNL. CONCLUSIONS: Partial peripheral nerve injury increases local CCL3 levels along the degenerating axons during Wallerian degeneration. This CCL3 binds to its receptor, CCR1, located on adjacent uninjured afferents, presumably nociceptors, to induce hyperalgesia in the neuropathic pain state.

Spinal and Peripheral GABA-A and B Receptor Agonists for the Alleviation of Mechanical Hypersensitivity following Compressive Nerve Injury in the Rat / 대한통증학회지

BACKGROUND: This study was conducted to investigate the roles of the spinal and peripheral gamma-aminobutyric acid (GABA)-ergic systems for the mechanical hypersensitivity produced by chronic compression of the dorsal root ganglion (CCD). METHODS: CCD was performed at the left 5th lumbar dorsal root ganglion. The paw withdrawal threshold (PWT) to von Frey stimuli was measured. The mechanical responsiveness of the lumbar dorsal horn neurons was examined. GABAergic drugs were delivered with intrathecal (i.t.) or intraplantar (i.pl.) injection or by topical application onto the spinal cord. RESULTS: CCD produced mechanical hypersensitivity, which was evidenced by the decrease of the PWT, and it lasting for 10 weeks. For the rats showing mechanical hypersensitivity, the mechanical responsiveness of the lumbar dorsal horn neurons was enhanced. A similar increase was observed with the normal lumbar dorsal horn neurons when the GABA-A receptor antagonist bicuculline was topically applied. An i.t. injection of GABA-A or GABA-B receptor agonist, muscimol or baclofen, alleviated the CCD-induced hypersensitivity. Topical application of same drugs attenuated the CCD-induced enhanced mechanical responsiveness of the lumbar dorsal horn neurons. CCD-induced hypersensitivity was also improved by low-dose muscimol applied (i.pl.) into the affected hind paw, whereas no effects could be observed with high-dose muscimol or baclofen. CONCLUSIONS: The results suggest that the neuropathic pain associated with compression of the dorsal root ganglion is caused by hyperexcitability of the dorsal horn neurons due to a loss of spinal GABAergic inhibition. Peripheral application of low-dose GABA-A receptor agonist can be useful to treat this pain.

Role of Peripheral Glutamate Receptors to Mechanical Hyperalgesia following Nerve Injury or Antidromic Stimulation of L5 Spinal Nerve in Rats with the Previous L5 Dorsal Rhizotomy / 대한통증학회지

BACKGROUND: Peripheral nerve injury leads to neuropathic pain, including mechanical hyperalgesia (MH). Nerve discharges produced by an injury to the primary afferents cause the release of glutamate from both central and peripheral terminals. While the role of centrally released glutamate in MH has been well studied, relatively little is known about its peripheral role. This study was carried out to determine if the peripherally conducting nerve impulses and peripheral glutamate receptors contribute to the generation of neuropathic pain. METHODS: Rats that had previously received a left L5 dorsal rhizotomy were subjected to a spinal nerve lesion (SNL) or brief electrical stimulation (ES, 4 Hz pulses for 5 min) of the left L5 spinal nerve. The paw withdrawal threshold (PWT) to von Frey filaments was measured. The effects of an intraplantar (i.pl.) injection of a glutamate receptor (GluR) antagonist or agonist on the changes in the SNL- or ES-produced PWT was investigated. RESULTS: SNL produced MH, as evidenced by decrease in the PWT, which lasted for more than 42 days. ES also produced MH lasting for 7 days. MK-801 (NMDAR antagonist), DL-AP3 (group-I mGluR antagonist), and APDC (group-II mGluR agonist) delayed the onset of MH when an i.pl. injection was given before SNL. The same application blocked the onset of ES-induced MH. NBQX (AMPA receptor antagonist) had no effect on either the SNL- or ES-induced onset of MH. When drugs were given after SNL or ES, MK-801 reversed the MH, whereas NBQX, DL-AP3, and APDC had no effect. CONCLUSIONS: Peripherally conducting impulses play an important role in the generation of neuropathic pain, which is mediated by the peripheral glutamate receptors.

Testosterone Causes Simultaneous Decrease of Ca2+i and Tension in Rabbit Coronary Arteries: by Opening Voltage Dependent Potassium Channels

The relationship between the level of testosterone and the incidence of coronary heart disease is still controversial in the view of the results of clinical and epidemiologic studies. This uncertainty might be partly due to relatively small number of experimental studies undertaken to investigate the cellular mechanism underlying the vascular responses to testosterone. To further investigate the cellular mechanisms of testosterone with respect to vascular response, we investigated the effect of testosterone on contractility and intracellular Ca2+ regulation in a rabbit coronary artery and evaluated the underlying mechanism of testosterone-induced changes of coronary vascular tone by using various pharmacological blockers. Testosterone was found to relax rabbit coronary arteries in a dose-dependent manner, and no significant difference was found in the relaxation response to testosterone with or without endothelium. Similar results were obtained in male and non-pregnant female rabbit coronary arteries. The relaxation response of rabbit coronary arteries to testosterone was greater for PGF2alpha-contracted rings than for KCl contracted rings, which suggest the involvement of K+ channels. Furthermore, the relaxation response to testosterone was significantly reduced by 4-aminopyridine, a sensitive blocker of voltage dependent K+ channels, but not by low doses of tetraethylammonium or iberiotoxin, a Ca2+ activated K+ channel blocker. Testosterone simultaneously reduced the intracellular Ca2+ concentration ([Ca2+]i) and tension, and 4-AP effectively antagonized the testosterone-induced change of [Ca2+]i and tension. Therefore, it may be concluded that the stimulation of voltage dependent K channels is responsible, at least in part, for the testosterone-induced relaxation of rabbit coronary arteries.

Loss of the Spinal GABAergic System Is Involved in Chronic Central Pain Following a Spinal Cord Injury; Behavioral and Electrophysiological Evidences / 대한마취과학회지

BACKGROUND: Allodynia, hyperalgesia, and spontaneous pain are symptoms characterized by chronic central pain which was frequently observed following a spinal cord injury (SCI). However, the underlying mechanism has not been fully understood. This study was conducted to investigate whether the loss of the GABAergic system in the spinal dorsal horn was involved in the development of central pain following a spinal cord injury. METHODS: SCI was induced by a hemisection of the spinal cord at T13 in adult male Sprague-Dawley rats. Mechanical allodynia was tested by measuring paw withdrawal frequency in response to repeated applications of a von Frey hair to the plantar surface of the hind-paw. Single neuronal activity of the dorsal horn neurons (L4 L6) was recorded extracellularly using a carbon filament-filled glass microelectrode (2 4 MOhm). The drugs were intrathecally or topically administrated on the spinal surface for behavioral and electrophysiological experiments, respectively. RESULTS: After a left spinal hemisection at T13, behavioral signs of mechanical allodynia developed on both hind-paws and responsiveness of spinal dorsal horn neurons increased on both sides of the spinal dorsal horn. GABA receptor agonists including GABAA and GABAB receptor subtypes suppressed mechanical allodynia on both sides of hind-paws and decreased responsiveness of spinal dorsal horn neurons on both sides of spinal cord. CONCLUSIONS: These results indicate that a loss of the GABAergic system within the spinal cord plays a key role on the development of central pain following a spinal cord injury.

Differentiation of neuroepithelial progenitor cells implanted into newborn rat brain striatum

It has been demonstrated that multipotent neuronal progenitor cells can be isolated from the developing or adult CNS and proliferated in vitro in response to epidermal growth factor. The present study was undertaken to investigate the differentiation of neuronal progenitor cells after transplantation into the neonatal rat forebrain striatum. Primary cultured progenitor cells were labeled with 3,3'-dioctadecycloxacarbonyl- amine perchlorate (DiO). DiO labeled progenitor cells were implanted into neonatal rat striatum. Implanted DiO labeled progenitor cells were differentiated into astrocytes and GABAergic neurons. These results suggest that implanted progenitor cells can be differentiated into neurons in host forebrain striatum. In addition, our data show that DiO labeling is a useful technique for tracing implanted progenitor cells.

Effects of iontophoretically applied substance P, calcitonin gene-related peptide on excitability of dorsal horn neurones in rats

Spontaneous pain, allodynia and hyperalgesia are well known phenomena following peripheral nerve or tissue injury, and it is speculated that secondary hyperalgesia and allodynia, are generally thought to depend on a hyperexcitability (sensitization) of neurons in the dorsal horn. It is supposed that the sensitization may be due to various actions of neurotransmitters (SP, CGRP, excitatory amino acids) released from the primary afferent fibers. In this study, we examined effects of the iontophoretically applied SP and CGRP on the response to EAA receptor agonists (NMDA and non-NMDA) in the WDR dorsal horn neurones and see if the effects of SP or CGRP mimic the characteristic response pattern known in various pain models. The main results are summarized as follows: 1) SP specifically potentiated NMDA response. 2) CGRP non-specifically potentiated both NMDA and AMPA responses. Potentiation of NMDA response, however, was significantly greater than that of AMPA response. 3) 50% of SP applied cells and 15.8% of CGRP applied cells showed reciprocal changes(potentiation of NMDA response and suppression of AMPA response). These results are generally consistent with the sensitization characteristics in diverse pain models and suggests that the modulatory effects of SP and CGRP on NMDA and non-NMDA (AMPA) response are, at least in part, contribute to the development of sensitization in various pain models.

The effect of carbon monoxide on contraction, cytosolic Ca2+ level and ionic currents in guinea pig ileal smooth muscle

The aim of this study was to clarify the mechanism of the inhibitory action of carbon monoxide (CO) on contraction, by measuring cytosolic Ca2+ level ((Ca2+)i) and ionic currents in guinea-pig ileum. CO (10%) inhibited 40 mM KCl-induced contraction and this effect was blocked by ODQ (1 micrometer), a soluble guanylyl cyclase (sGC) inhibitor. CO inhibited the 40 mM KCl-induced contraction without changing (Ca2+)i. Cumulative addition of KCl induced a graded increase in (Ca2+)i and muscle tension. In the presence of CO, cumulative addition of KCl induced smaller contraction than in the absence of CO. On the other hand, the increase in (Ca2+)i induced by cumulative addition of KCl was only slightly decreased in the presence of CO, and the (Ca2+)i-tension relationship shifted downwards. Using the patch clamp technique with a holding potential of -60 mV, we found that CO had little effect on the peak Ba currents (IBa) when voltage was stepped from -60 mV to 0 mV. In addition, CO showed no effect on the depolarization-activated outward K+ currents in the all potential ranges. We conclude that CO inhibits smooth muscle contraction mainly by decreasing the Ca2+ sensitivity of contractile elements via a cGMP-dependent pathway, not by involving L-type Ca2+ and outward-potassium currents in guinea-pig ileum.

Adrenergic sensitivity of uninjured C-fiber nociceptors in neuropathic rats

We investigated the adrenergic sensitivity of afferent fibers in the L4 dorsal roots of rats with a unilateral ligation of the L5-L6 spinal nerves. About 12% of nociceptive fibers on the affected side were excited by sympathetic stimulation or by intra-arterial injection of norepinephrine which did not affect A beta-fiber activity. Sympathetic excitation of nociceptive fibers was suppressed by alpha 1-antagonist prazosin, while it was unaffected by alpha 2-antagonist yohimbine. Most of these fibers were excited by intra-arterial injection of alpha 1-agonist phenylephrine, without being affected by an injection of alpha 2-agonist clonidine. Sympathetic excitation was blocked by lidocaine applied near the receptive fields of recorded fibers. The results suggested that some nociceptors remaining intact after partial nerve injury become sensitive to sympathetic activity by the mediation of alpha 1-adrenoceptors in the peripheral endings.

Transcutaneous Electrical Nerve Stimulation and Microcurrent Electrical Neuromuscular Stimulation on Pain Behaviors in Rats with an Experimental Neuropathy

OBJECTIVE: The present study was conducted to investigate the effects of transcutaneous electrical nerve stimulation (TENS) and microcurrent electrical neuromuscular stimulation (MENS) on pain-like behaviors developed in rats with an experimental neuropathy. METHOD: Neuropathic surgery was done by a unilateral ligation of L5 and L6 spinal nerves of the rat. Allodynic behavior was examined by measuring foot withdrawal frequency in response to 10 applications of a von Frey filament (2.5 g) to the plantar surface of the foot. Ongoing pain behavior was examined by measuring cumulative time in 3 min that the rat lifted its foot off a plate held at cold temperature (5oC). TENS (square pulses; 3 Hz, 30 mA) or MENS (bipolar pulses; 10 Hz, 300 microampere) was applied for 15 min or 5 min, respectively, to the skin of the affected foot. RESULTS: Behavioral signs of mechanical allodynia and cold-induced ongoing pain had developed after nerve injury. Either TENS or MENS, when applied once, alleviated allodynic behavior, lasting up to 2 hrs. Such an alleviation lasted much longer when TENS or MENS was applied repeatedly (once a day for 6 days); 3 days by TENS and 1 day by MENS. Cold-induced ongoing pain behavior, however, was not affected by the repeated application of either TENS or MENS. CONCLUSION: The results suggest that both TENS and MENS are useful tools for the treatment of mechanical allodynia. Repeated application of TENS or MENS is more effective in alleviating mechanical allodynia than its single application. Either TENS or MENS, however, seems not effective in alleviating cold-induced ongoing pain.

Effects of iontophoretically applied naloxone, picrotoxin and strychnine on dorsal horn neuron activities treated with high frequency conditioning stimulation in cats

Transcutaneous electrical nerve stimulation(TENS), acupuncture-needling, and electroacupuncture are useful non-ablative methods in medical practice for relief of pain. These procedures appear to work by causing an increased discharge in afferent nerve fibers which in turn modifies the transmission of impulses in pain pathways. It is known that the mechanism of analagesic effect via these maneuvers are variable depending on the stimulating parameters. For example, the endogenous opioid system is profoundly related to the mechanism when a peripheral nerve stimulation is applied with parameters of low frequency and high intensity. However, when stimulated with parameters of high frequency and high intensity, the reduced activity of dorsal horn neurons is only slightly reversed by a systemic administration of naloxone, a specific opiate antagonist. Thus, the present study was performed to investigate the neurotransmitter that concerns the mechanism of peripheral nerve stimulation with parameters of high frequency and high intensity. We used an iontophoretic application of antagonists of possible related neurotransmitters. The dorsal horn neuron activity which was evoked by squeezing the peripheral cutaneous receptive field, was recorded as an index of pain with a microelectrode at the lumbo-sacral spinal cord. Naloxone, picrotoxin and strychnine were applied at 200nA during a period of conditioning nerve stimulation. We observed the effects of these drugs on the change of dorsal horn neuron activities. The main results of the experiment can be summarized as follows. The spontaneous activity of dorsal horn neurons increased in the presence of glutamate and decreased with GABA. It did not change with naloxone, picrotoxin or strychnine. When naloxone was applied iontophoretically during peripheral nerve stimulation, there was no statistically significant analgesic effect compared with that of the control group. When picrotoxin was applied iontophoretically during peripheral nerve stimulation, the analgesic effect was reduced. When strychnine was applied, the analgesic effect was reduced but did not show a statistically significant difference with the control group. These results suggested that the GABAergic system may have been partially related in the analgesic action of peripheral nerve stimulation with parameters of high frequency and high intensity.

Mechanism of transmission and modulation of renal pain in cats; effect of nucleus raphe magnus stimulation on renal pain

Initially, when periaqueductal gray (PAG) is electrically stimulated, analgesia is induced, and this phenomenon is called stimulation-produced analgesia. Nucleus raphe magnus (NRM) as well as PAG are known to be the potent analgesic centers. NRM could modulate the nociceptive response of spinal cord neurons through spinally projecting fibers. However, as well as the above analgesic effects have been confined to the somatic pain, it was variable according to species, and the analgesic effect by NRM stimulation on the visceral pain was not yet clarified. In this study the analgesic effect by NRM stimulation on the visceral pain was examined through recording the activities of the dorsal horn neurons with renal input and renal pain, as a type of visceral pain. The renal pain was induced by ureteral occlusion or renal arterial occlusion, which in turn activated the renal mechanoreceptor or chemoreceptor. These cells had concomitant somatic input. In order to compare the effects of NRM stimulation on the renal pain with somatic pain, the somatic stimulation such as squeezing was conducted on the peripheral receptive field. The main results are summarized as follows: 1) After an electrical stimulation of NRM, spontaneous activities of dorsal horn neurons with renal input were reduced to 73.3 +/- 9.7% of the control value. 2) After an electrical stimulation of NRM, activities of dorsal horn neurons with renal input evoked by a brush, a type of non-noxious stimuli, did not change significantly. But the activities by a squeeze, a type of noxious stimuli, the activities were reduced to 63.2 +/- 7.2% of the control value. 3) After an electrical stimulation of NRM, activities of dorsal horn neurons with renal input evoked by occlusion of ureter or renal artery were reduced to 46.7 +/- 8.8% and 49.0 +/- 8.0% of the control value respectively. 4) The inhibitory effect of NRM on the dorsal horn neurons with renal input did not show any difference between renal A delta fiber and C fiber group. 5) By the electrical stimulation of NRM, the activities evoked by ureteral occlusion showed more reduction in the high threshold cell group than in the wide dynamic range cell group. These results suggest that activation of NRM can alleviate the renal pain as well as the somatic pain by modulating the dorsal horn neurons activities.

Mechanism of transmission and modulation of renal pain in cats: effects of transcutaneous electrical nerve stimulation on renal pain

Transcutaneous electrical nerve stimulation (TENS) has widely been employed as a method of obtaining analgesia in medical practice. The mechanisms of pain relief by TENS are known to be associated with the spinal gate control mechanism or descending pain inhibitory system. However, most of the studies concerning the analgesic effects and their mechanisms for TENS have dealt with somatic pain. Thus, in this experiment, we investigated the analgesic effects of TENS on renal pain as a model of visceral pain, and the characteristics of the dorsal horn cells with renal inputs. The renal pain was induced by acute occlusion of the ureter or renal artery. The main results are summarized as follows: 1) The renal nerve was composed of A beta, A delta and C fiber groups; the thresholds for each group were 400-800 mV, 1.1-1.5 V, and 2.1-5.8 V, respectively. 2) The dorsal horn cells tested received A and/or C afferent fibers from the kidney, and the more C inputs the dorsal horn cells had, the greater was the response to the stimuli that elicited the renal pain. 3) 94.9% of cells with renal input had the concomitant somatic receptive fields on the skin; the high threshold (HT) and wide dynamic range (WDR) cells exhibited a greater responses than low threshold (LT) cells to the renal pain-producing stimuli. 4) TENS reduced the C-responses of dorsal horn cells to 38.9 +/- 8.4% of the control value and the effect lasted for 10 min after the cessation of TENS. 5) By TENS, the responses evoked by acute occlusion of the ureter or renal artery were reduced to 37.5 +/- 9.7% and 46.3 +/- 8.9% of the control value, respectively. This analgesic effects lasted 10 min after TENS. 6) The responses elicited by squeezing the receptive fields of the skin were reduced to 40.7 +/- 7.9% of the control value and the effects lasted 15 min after TENS. These results suggest that most of dorsal horn cells with renal inputs have the concomitant somatic inputs and TENS can alleviate the renal pain as well as somatic pain.

Effect of peripheral nerve stimulation on the dorsal horn cell activity in cats with cutaneous inflammation

There are some reports showing that an experience of long-enduring pain causes a change in the pain transmission system, suggesting a plastic nature of the nociceptive system. However, most of the studies concerning the analgesic effect of peripheral nerve stimulation dealt with normal animal or human subjects. So, the present study was undertaken to investigate the effect of peripheral nerve stimulation on the dorsal horn cell activity using a tonic pain model, which was made by producing a cutaneous inflammation. The main results are summarized as follows. 1) The evoked activity by electrical or natural stimulation as well as spontaneous activity was enhanced, and the receptive field size was also expanded by the inflammation. 2) Peripheral nerve conditioning stimulation reduced the C-response of the dorsal horn cell in the normal and inflamed group, and the degree of inhibition between the two groups showed no significant difference. 3) Inhibition of the C-response of the dorsal horn cells by peripheral conditioning stimulation was completely reversed by naloxone in the inflamed group whereas there was a partial block in the normal group.

The Effect of D-phenylalanine on the Analgesia Produced by Peripheral Nerve Conditioning Stimulation in the Cat

An effect of D-phenylalanine on the pain inhibitory mechanism of prolonged electrical stimulation of the peripheral nerve was studied in decerebrate cats and spinal cats. The response of spinal neurons was elicited either by electrical stimulation of the ipsilateral common peroneal nerve and tibial nerve. The single-unit activity of motor neurons which represent the flexion reflex was recorded from a filament of ventral rootlet divided from either the L7, S1 or S2 ventral root, and activity of dorsal horns cells was recorded with a microelectrode at the lumbosacral cord The conditioning stimuli which provocate the pain inhibitory mechanism of the common peroneal or tibial nerve was applied with repetitive, low frequency (2Hz), at a suprathreshold intensity for C fiber, for 30-45 minutes. The results of the experiment are summarized as follows: 1. Applying conditioning stimuli produced a powerful inhibition of the responses which was provocated by noxious stimuli in either the decerebrate or the spinal cat without any statistical difference, and this effect can be observed for 15 minutes after the cessation of the conditioning stlmuli 2. This response was reversed completely by systemic injection of a specific opiate antagonist, naloxone. It suggests that the conditioning stimulus of the peripheral nerve can produce the endogenous opiate related pain inhibitory effect as the spinal mechanism. 3. The conditioning stimuli can produce the analgesic effect by means of supression of the activity of the dorsal horn cell which was related to the pain response in the decerebrate cat. The same result could be observed in flexion reflex. 4. D-phenylalanine, a putative inhibitor of carboxypeptidase which degradates the endogenous opiate-enkephalin, was studied in this experiment under the hypothesis that D-phenylalanine will emphasize or prolongate the action of enkephalin. But, intravenously injected D-phenylalanine did not potentiate the inhibitory effect of the conditioning stimuli of the peripheral nerve. From the above result, it is speculated that the electrical stimulation of the peripheral nerve is directly mediated by an endogenous opiate related analgesia, and the site of the analgesic action resides mainly in spinal cord level. But these data could not support the gypothesis that antinociceptive effect of D-phenylalanine results frm the potentiation of endogenously released enkephalin.

Relationship Between Dorsal Horn Cell Activity and Electrical Stimulation of Peripheral Nerve with Special Reference of Stimulatory Parameters

Transcutaneous electrical nerve stimulation (TENS), acupuncture-needling, and electroacu! Puncture are useful non-ablative methods in medical practice for relief of acute and chronic r pain These procedures appear to work by causing an increased discharge in afferent nerve fibers which in turn modifies the transmission of impulses in pain pathways. The present study was performed to evaluate the analgesic effects of peripheral nerve stimulation with different stimulatory parameters in decerebrated cats and spinalized cats. And we studied the effects of naloxone, a specific opiate antagonist, on analgesia produced by 50 Hz, C intensity conditioning stimulation. The electrical response of.spinal neurons was elicited either by electrical stimulation of the ipsilateral common peroneal nerve or tibial nerve, and then the single unit activity of the dorsal horn cell was recorded with a carbon filament-filled glass microelectrode at the lumbosacral spinal cord. The conditioning stimuli which provoke the pain inhibitory mechanism were applied to the cornmon peroneal nerve or tibial nerve with a relatively high frequency (25, 50, 200Hz) for 15, 30, and 60 seconds at suprathreshold intensity for A delta or C fiber. The results of the experiment are summarized as follows: 1. Peripheral conditioning stimulation at C strength showed larger analgesic effects than those produced by stimulation at A delta strength. And analgesic effects produced by conditioning stimulation for 30sec were greater than those produced by stimulation for 15sec, but showed no statistically significant difference from those produced by stimulation for 60 sec. 2. Analgesic effects produced by 50Hz conditioning stimulation were greater than thoseproduced by 25Hz stimulation. But 200Hz stimulation showed a lesser analgesic effect than 50 or 25Hz conditioning stimulation. 3. The analgesic effect produced by 50Hz conditioning stimulation was only slightly affected by naloxone, a specific opiate antagonist, indicating that involvement of an endogenous opiate system was minimal. 4. The analgesic effect produced by conditioning stimulation in decerebrated cats was nearly the same as in spinal cats suggesting that the neural circuitry responsible for the analgesic action seems to reside mostly within the spinal cord. From the above results, it is concluded that 1) frequency of stimulation is important for an efficient analgesia, i.e., stimulation with excessively high frequency decreases the analgesic effect, 2) the analgesic effect produced by high frequency conditioning stimulation may be minimally mediated by an endogenous opiate system, and 3) the site of analgesic action resides mainly in the spinal cord.

Effects of ketamine on contractile responses in vascular smooth muscle

This study was designed to determine the effects of ketamine on contractions induced by norepinephrine (NE), K+ or histamine (Hist) and on agonist-induced calcium mobilization, in rabbit thoracic aorta with or without endothelium. Contractile responses to NE, K+ or Hist were markedly attenuated by prior exposure to ketamine. Subsequent addition of ketamine to the rabbit aorta undergoing an isometric contraction induced by NE, K+ or Hist also decreased the contractile responses in a calcium ion concentration-dependent manner. Preincubation with ketamine produced a concentration-dependent inhibition of contractile responses elicited by the addition of calcium ion (1.6 mM) to a Ca(++)-free depolarizing solution. However, the phasic contraction produced by NE with 2mM lanthanum pretreatment, which is release of intracellular calcium, was also inhibited by ketamine. Moreover, the tonic contraction produced by NE after depletion of the agonist-releasable pool of intracellular calcium, which is thought to be due to calcium influx, was depressed by ketamine. These data suggest that ketamine relaxes NE-contracted rings of rabbit thoracic aorta by decreasing calcium entry and by producing an extracellular calcium-independent relaxant effect.

Effect of Caffeine on Calcium Flux across the Sarcolemma of Guinea Pig Atrial Trabeculae during Contracture

The changes in extracellular calcium activities during contractures of guinea pig atrial trabecular muscles were measured with Ca2+-selective electrodes. When the tissue was superfused with Na+-free Tyrode solution extracellular Ca2+ activities were decreased and contractures were induced with some delay. When the contracture was relaxed with Na+-containing Tyrode solution, extracellular Ca2+ activities were increased transiently and recovered in a Na+-dependent manner. The magnitude of extracellular Ca2+ activity decreased was proportional to the maximum magnitude of contracture induced by Na+-free solution. Addition of caffeine (10 mM) to Na+-free solution induced transient contracture following slow development of contracture and an increase in extracellular Ca2+ activity. Removal of caffeine from Na+-free solution caused a slow relaxation of contracture and a decrease in extracellular Ca2+ activity. These results confirm that caffeine blocks Ca2+ uptake by the sarcoplasmic reticulum (SR) resulting in an increase in sarcoplasmic Ca2+ activity. Ca2+ activity in the extracellular space, the amount of Ca2+ transported into the cell(Ca2+ depletion in the extracellular space), and the magnitude of contracture are well correlated. Present experiments suggest that extracellular use of Ca2+-selective electrodes provides continuous and quantitative monitoring of Na+-dependent Ca2+ flux across the cardiac cell membrane.