Characterization of 5-HT Receptors in Rat Proximal Colon

• 1.In view of its multiple sites of action, we investigated the activity of 5-hydroxytryptamine (5-HT, serotonin) on various potential receptors in the isolated proximal colon of rats.
• 2.5-HT induced concentration-dependent contractions of colonic strips (pEC₅₀=7.54±0.12).
• 3.The 5-HT₁ receptor agonist, 5-carboxamidotryptamine, induced concentration-dependent contractions (pEC₅₀=5.93±0.27); however, neither the 5-HT₃ receptor-agonist, phenylbiguanide, nor the 5-HT₄ receptor-agonist, renzapride, caused contractions at concentrations as high as 10⁻⁴ M.
• 4.The 5-HT_1/2 receptor antagonist, methiothepin, caused concentraction-dependent nonsurmountable antagonism. The 5-HT₃ receptor antagonist, tropisetron, inhibited the contractions to a concentration of 5-HT⩾10⁻⁶ M. Ketanserin had no effect on responses to 5-HT.
• 5.Tetrodotoxin and atropine had no effect on responses to 5-HT.
• 6.We conclude that contractions to 5-HT are mediated by 5-HT_1-like and probably 5-HT₃ receptors that activate tetrodotoxin insensitive mechanisms.

     

5-HT receptors on identified Lymnaea neurones in culture: Pharmacological characterization of 5-HT3 receptors

• 1.1. The selective agonist, 1-(m-chlorophenyl)-biguanide (m-CPBG) and antagonist, 3-tropanyl-3,5-dichlorobenzoate (MDL 72222) were used to characterize the 5-HT₃ receptors in cultured identified neurones; the serotonin-containing cerebral giant cells (CGCs) and some follower neurones in the buccal ganglia of Lymnaea stagnalis.
• 2.2. 5-HT and its agonists were pressure ejected, while the 5-HT antagonists were bath applied.
• 3.3. Although m-CPBG evoked mostly depolarizing responses, hyperpolarizing responses were sometimes evoked.
• 4.4. At 10⁻⁴ M, m-CPBG failed to mimic the responses of 5-HT, but at a concentration higher. 10⁻³ M, pressure-ejected m-CPBG mimicked most 5-HT responses.
• 5.5. The 5-HT₂ antagonist ketanserin failed to block the m-CPBG-evoked responses, whilst partially blocking the 5-HT responses.
• 6.6. These results suggest the presence of 5-HT₃ receptors similar to those found in mammalian neurones, and that multiple subtypes of these receptors may be present in Lymnaea neurones.

     

A lack of supersensitivity to opioid receptor agonists following chronic spinal opioid receptor antagonist administration in the rat

• 1.1. Male Sprague-Dawley rats were chronically tested with intrathecal (i.t.) receptor selective opioid antagonists to determine if antinociceptive supersensitivity developed to selective i.t. opioid receptor agonists.
• 2.2. A subcutaneously implanted osmotic minipump was used to deliver the μ-opioid receptor antagonist CTOP (0.3 nmol) or the b-opioid receptor antagonist naltrindole (5.5 nmol) for 7 days.
• 3.3. Following a 24 hr washout period, rats received a single i.t. dose (ED₅₀) of either DAMPGO (for CTOP-treated animals) or DPDPE (for naltrindole-treated animals) and the antinociceptive effects of the agents were tested on the tail-flick test.
• 4.4. Our findings revealed that chronic spinal treatment with selective opioid receptor antagonists did not induce an antinociceptive supersensitivity to selective opioid receptor agonists.
• 5.5. Perhaps this lack of supersensitivity is reflective of difficulties inherent to opioid receptor antagonists that do not possess negative intrinsic activity

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Pharmacological analysis of the vasodepressor effect of KRN2391 in pithed rats

• 1.1. The antagonism by glibenclamide of the vasodepressor effects of KRN2391, cromakalim and nitroglycerin was compared in pithed rats with blood pressure supported by an infusion of phenylephrine.
• 2.2. Cumulative administration of KRN2391 (3–300 μg/kg, i.v.), cromakalim (3–300 μg/kg, i.v.) and nitroglycerin (1–300 μg/kg, i.v.) produced dose-dependent decreases in diastolic blood pressure.
• 3.3. In rats given glibenclamide (20 mg/kg, i.v.), the dose-vasodepressor curves for KRN2391 and cromakalim were shifted to the right. However, glibenclamide had no effect on the vasodepressor effect of nitroglycerin.
• 4.4. The ED_{50 mmHg} values increased about 5.9 fold for KRN2391 and 9.5 fold for cromakalim in glibenclamide-treated rats.
• 5.5. These results suggest that the vasodepressor effect of KRN2391 is due to both glibenclamide-sensitive and insensitive mechanisms. This glibenclamide-insensitive effect of KRN2391 is thought to reflect its nitrate action.

     

Effects of diltiazem and verapamil on ADP-induced rabbit platelet shape change and aggregation

• 1.1. The effects of diltiazem and verapamil (two structurally different calcium channel blockers) were examined on the rabbit platelets shape change and aggregation induced by adenosine-5′-diphosphate (ADP).
• 2.2. ADP was a much more potent stimulator on inducing platelet shape change (ED₅₀ = 1 × 10⁻⁷) than platelet aggregation (ED₅₀ = 1.78 × 10⁻⁶).
• 3.3. Both drugs similarly inhibited ADP-induced platelet shape change and aggregation at concentrations more than 300 μM.
• 4.4. There were no significant differences in inhibitory effects of either diltiazem or verapamil on ADP-induced platelet shape change and aggregation.
• 5.5. The inhibitory effects of diltiazem and verapamil on ADP-induced platelet shape change and aggregation at high concentrations may be due to their non specific properties.

     

Pre- and postjunctional effects of 5-hydroxytryptamine in dog cephalic veins of neonates

• 1.1. This investigation was undertaken to determine the existence of pre- and post junctional effects of 5-hydroxytryptamine (5-HT) in the cephalic vein of newborn dogs. For the sake of comparison, some experiments were also carried out on the veins of adult animals.
• 2.2. 5-HT reduced, in a concentration-dependent manner the overflow of tritium evoked by electrical stimulation of tissues previously loaded with ³H-noradrenaline for the same range of concentrations (0.01–1μM 5-HT) in both neonates and adults. The maximal reduction of tritium overflow caused by 5-HT was 36±6% and 77±5% (n=4; P<0.001) in neonates and adults, respectively.
• 3.3. Postjunctionally, 5-HT caused concentration-dependent contractions of vessels from both neonates and adults and its EC₅₀ values were not significantly different at the two ages. The maximal effect of 5-HT was smaller in newborn (0.69±0.03 g/mg) than in adult (1.16±0.06 g/mg) animals (n=9; P<0.01).
• 4.4. Metitepin antagonized 5-HT effects while ketanserin was ineffective, both pre- and postjunctionally, in newborn and adult dogs.
• 5.5. It is concluded that there are 5-HT receptors at pre- and postjunctional level at birth and that these receptors probably belong to the 5-HT₁ type.

     

LiCl uncouples signal transduction in morphine-induced supraspinal antinociception in mice

• 1.1. The present study examined whether LiCl antagonism of morphine-induced antinociception in mice occurs at μ-opioid receptors.
• 2.2. The antinociceptive ED₅₀ value of intracerebroventricular morphine was maximally increased compared to controls 18 hr after LiCl (10 mmol/kg, s.c.) and remained significantly less (P < 0.05) 7 and 14 days after once-daily LiCl treatment.
• 3.3. There was no significant difference in [³h]-[d-Ala²,N-MePhe⁴,Gly-ol⁵]enkephalin affinity or receptor density compared to controls (K_D = 0.43 nM, B_max = 54.8 ± 9.3 pM).
• 4.4. These results suggest that LiCl's effect is not on μ-opioid receptors, but rather on some distal site.

     

Increased toxicity of methamphetamine in morphine-dependent mice

• 1.1. The effect of methamphetamine on morphine-dependent mice was investigated by calculating the LD₅₀ (IP), measuring motor activity, anorectic actions, and body temperature.
• 2.2. Methamphetamine was more toxic in morphine-dependent mice (LD₅₀=20.6 mg/kg) than in normal mice (LD₅₀=43.2 mg/kg).
• 3.3. Methamphetamine-induced locomotor activity was greater in morphinized than in nonmorphinized mice at doses of 2.5 and 5 mg/kg IP.
• 4.4. Methamphetamine also increased the body temperature of morphinized mice more than that of normal mice (P<0.05).
• 5.5. These findings suggest that methamphetamine is more toxic in morphine-dependent than in nondependent mice.

     

Role of calcium in mediating the biphasic contraction of the rabbit urinary bladder

• 1.1. The response of the urinary bladder to field stimulation is biphasic in nature consisting of an initial phasic contraction followed by a prolonged tonic phase which lasts for the duration of the stimulation.
• 2.2. The phasic response is mediated by the release of neurohumoral transmitters, primarily acetylcholine (via muscarinic receptor stimulation) and ATP (via purinergic receptor stimulation). The tonic component is mediated entirely via muscarinic receptor stimulation.
• 3.3. The present study investigates the dependence on extracellular calcium of the phasic and tonic contractile responses to field stimulation, bethanechol, and ATP. The results can be summarized as follows:
• 4.4. Field stimulation (2 and 32 Hz) and bethanechol evoke a biphasic contractile response whereas ATP evokes only a phasic response.
• 5.5. There were no significant effects of either calcium channel blockers or calcium fee EGTA medium on either spontaneous contraction or basal tension of muscle strips.
• 6.6. The calcium channel antagonists diltiazem and verapamil inhibited both the phasic and tonic responses induced by field stimulation (both 2 and 32 Hz) in a dose dependent manner.
• 7.7. For both 2 and 32 Hz stimulation, the ED₅₀s for the inhibition of the tonic phases of the responses to field stimulation were significantly lower than the ED₅₀s for the inhibition of the phasic responses.
• 8.8. The tonic phase of the responses to field stimulation were inhibited to a significantly greater degree than the phasic responses by incubation in calcium-free medium containing EGTA.
• 9.9. Both the phasic and tonic components of the response to bethanechol stimulation were inhibited equally, and followed a similar time course as the tonic component of field stimulation.
• 10.10. The response to ATP was relatively insensitive to calcium depletion.
• 11.11. In summary, the tonic response to field stimulation (purely cholinergic) is significantly more dependent on extracellular calcium than is the phasic response (cholinergic + purinergic).

     

Investigation into the effect of 5-hydroxytryptamine on fluid transport in the rat small intestine

• 1.1. 5-Hydroxytryptamine (5-HT) has been shown to cause a consistent secretory effect in the rat small intestine only when administered luminally or by close intraarterial infusion. Intraluminal 5-HT-induced secretion is possibly mediated by 5-HT₄ receptors. Therefore, it was decided to investigate the effect of 5-HT and selective 5-HT₄ receptor agonists (SC 53116 and DAU 6236) on intestinal fluid transport in rat jejunum and ileum. The study also investigated the effect of a selective 5-HT₄ receptor antagonist (GR 113808) against the intraluminally administered 5-HT.
• 2.2. 5-HT receptor agonists and antagonists were administered intraluminally in pentobarbitoneanesthetized rats. Changes in intestinal fluid transport across the intestinal wall were measured by a single pass technique.
• 3.3. Intraluminal 5-HT produced significant antiabsorptive effects is both the jejunum and ileum. The 5-HT-induced responses were blocked by intraluminal administration of the 5-HT₄ receptor antagonist GR 113808. The 5-HT₄ agonist SC 53116 induced antiabsorptive effects in both regions of the small intestine, but DAU 6236 did not affect the rates of fluid transport.
• 4.4. The results indicate that a 5-HT₄ receptor has a role in the luminal 5-HT-induced antiabsorptive effect on intestinal fluid transport in the rat.

     

Sodium nonivamide acetate: A non-pungently antinociceptive capsaicin derivative with unusual anti-inflammatory properties

• 1.1. Bradykinin-induced vascular pain in conscious rats, hyperalgesia in the rat hind paw, rat hind paw edema induced by compound 48/80 and carrageenin and dye exudation induced by intraperitoneal injection of 0.7% acetic acid in mice were all inhibited by sodium nonivamide acetate (SNA).
• 2.2. Collagen and arachidonic acid-induced rabbit platelet aggregations were inhibited by SNA and capsaicin. In human platelet microsomes, prostaglandin E₂ formation in arachidonic acid metabolite was not inhibited by SNA but was inhibited by capsaicin and indomethacin; thromboxane B₂ formation and its synthetase activity were inhibited by SNA and capsaicin.
• 3.3. In the extracellular recording, SNA could not decrease the action potential amplitude of the vagus nerve.
• 4.4. The motor activity of mice induced by caffeine (1.0 mg/kg) was inhibited by SNA and capsaicin.

     

Inhibition of rats adjuvant arthritis by a bradykinin antagonist hoe 140 and its influence on kallikreins

• 1.1. This study examines the effect of Hoe 140, a bradykinin (BK) 2 receptor antagonist, indomethacin and prednisolone on chronic adjuvant arthritis of the knee in rats. We also evaluated the influence of Hoe 140 on BK-forming enzymes in the synovial and paw tissues.
• 2.2. Adjuvant arthritis was induced in male Sprague-Dawley rats in the right knee by injecting 0.05 ml of a fine suspension of heat-killed Mycobacterium tubercle bacilli in liquid paraffin (5 mg/ml).
• 3.3. Hoe 140 (1.5 mg/kg i.p.), indomethacin (2.5 mg/kg orally) and prednisolone (3.0 mg/kg orally) administration for 9 days resulted in significant suppression of knee joint swelling. Plasma and tissue kallikrein levels were raised (P<0.01) in the synovial and paw tissues of adjuvant arthritic rats. Hoe 140 treatment reduced (P<0.05) tissue kallikrein but increased (P<0.01) plasma kallikrein levels in synovial tissue.
• 4.4. Hoe 140 treatment did not alter (P>0.05) the raised plasma and tissue kallikrein levels in the paw tissue. The findings indicate that Hoe 140 may be a useful anti-inflammatory agent and BK plays a major role in this adjuvant-induced arthritis model.

     

Post-natal decrease in chronotropic sensitivity to acetylcholine in rat heart

• 1.1. The negative chronotropic effects of acetylcholine and carbachol on isolated rat right atria were examined at 0, 4, 8 and 16 weeks after birth.
• 2.2. Acetylcholine produced negative chronotropic responses at all ages and completely abolished spontaneous beating at its maximum effective concentration.
• 3.3. The sensitivity to acetylcholine, expressed in terms of ED₅₀ values, was higher at 0 and 4 weeks than at 8 and 16 weeks, ED₅₀ values (μM) at 0, 4, 8 and 16 weeks being 9.5 ± 1.8 (n = 12), 13.2 ± 3.4 (n = 11), 59.3 ± 10.9 (n = 14) and 51.5 ± 17.5 (n = 5), respectively.
• 4.4. Neostigmine produced a leftward shift of the concentration-response curve for acetylcholine both at 4 and 8 weeks after birth. The shift was larger at 8 weeks and no difference in sensitivity to acetylcholine was observed between the two ages in the presence of neostigmine.
• 5.5. Further, no developmental changes were observed in the sensitivity to carbachol, which is not hydrolyzed by cholinesterase.
• 6.6. We concluded that the chronotropic sensitivity to acetylcholine of rat atria decreases post-natally during the period between 4 and 8 weeks after birth due to increase in cholinesterase activity.

     

Different calcium dependencies of contractile activity of prostatic and epididymal portions of rat vas deferens

• 1.1. The effects of some organic calcium entry blockers and different concentrations of extracellular calcium on electrically-evoked contractions of isolated epididymal and prostatic portions of rat vas deferens were investigated.
• 2.2. Both epididymal and prostatic parts of rat vas deferens responded to single pulse or train electrical field stimulation, with twitch contractions of submaximal amplitude.
• 3.3. Verapamil showed a biphasic action on the contractions produced by single pulse electrical stimulation. In concentrations < 10⁻⁵M, it potentiated the responses of both portions, but at higher concentrations, the excitatory action was overcome by a concentration-dependent inhibitory effect.
• 4.4. Nifedipine reduced the amplitude of electrically-evoked contractions of both portions in a concentration-dependent manner. The ED₅₀ of nifedipine was 3.6 × 10⁻⁸M and 2.1 × 10⁻⁶M in prostatic and epididymal portions, respectively.
• 5.5. Dantrolene sodium reduced the amplitude of electrically-evoked contractions of both portions in a concentration-dependent manner. The ED₅₀ of dantrolene was 1.55 × 10⁻⁴M and 9.1 × 10⁻⁴M in prostatic and epididymal portions, respectively.
• 6.6. Reduction of Ca²⁺ concentration in medium reduced the amplitude of contractions of both portions significantly. This calcium dependence was more apparent in low frequencies of electrical stimulation.

     

Effects of tandospirone on second messenger systems and neurotransmitter release in the rat brain

• 1.1. We studied the effects of tandospirone, a novel serotonin (5-HT)_1A receptor-related anxiolytic, on the intracellular second messenger systems and neurotransmitter release.
• 2.2. Tandospirone inhibited forskolin-stimulated adenylate cyclase activity in rat hippocampal membranes by activation of 5-HT_1A receptors and had high efficacy comparable to 5-HT_1A receptor agonists such as 5-HT and 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT).
• 3.3. Tandospirone suppressed carbachol-stimulated phosphatidyl-inositol metabolism (PI response), which was shown to be a 5-HT_1A receptor-mediated event.
• 4.4. Tandospirone did not affect the release of 5-HT, norepinephrine (NE), dopamine (DA) and acetylcholine (ACh) from rat brain slice preparations.
• 5.5. These findings suggested that tandospirone shows high agonistic efficacy on the postsynaptic 5-HT_1A receptors but does not affect the presynaptic autoreceptors located on nerve endings. The modulation of the second messenger system via postsynaptic 5-HT_1A receptors might be involved in the anxiolytic efficacy of tandospirone.

     

Effects of the selective 5-HT receptor agonists 8-OHDPAT and DOI on behavior and brain biogenic amines of rats

• 1.1. The behavioral responses, as well as the biogenic amines and metabolite contents in discrete brain areas were determined in male rats subcutaneously treated with a 5-HT_1A (8-OHDPAT) or 5-HT_2A (DOI) agonist at doses (0.5-2 mg/kg) sufficient to produce the typical effects of the stimulation of these brain receptor subtypes.
• 2.2. Besides the expected effects (i.e., forepaw treading, flat body posture and inhibition of 5-HT release and turnover), 8-OHDPAT displayed signs of increased dopaminergic transmission.
• 3.3. DOI increased dopamine turnover and provoked stereotypical behavior, in addition to head shakes and body twitches.
• 4.4. Moreover, DOI induced both forepaw treading and flat body posture, which are believed to be typical responses to the stimulation of brain 5-HT_1A receptors.
• 5.5. This finding cannot be explained on the basis of actual knowledge, because the affinity of DOI for 5-HT_1A receptor has been found to be very low, whereas indirect mechanisms of activation of this receptor subtype triggered by stimulation of 5-HT_2A receptor are actually unknown.

     

Central and peripheral 5-HT receptors in the leech (Hirudo medicinalis) redefined

• 1.1. 5-Hydroxytryptamine (5-HT) receptors in the Retzius neurones (R cells) and longitudinal body wall muscle strips of the leech H. medicinalis were studied using a range of selective agonists developed for mammalian receptors.
• 2.2. All agonists induced hyperpolarisation of R cells: the order of their potency was 5-HT > 5-CT > 2-Me 5HT > α-Me-5-HT > > CGS-12066B=5MeOT > PAPP > Buspirone. This receptor is most like a mammalian 5-HT_1c/2 receptor.
• 3.3. 5-HT induces relaxation of body wall strips, preceded at higher doses by contraction or an increase in spontaneous contractions. The relaxing effect was best mimicked by 5-CT and α-Me-5-HT, suggesting a 5-HT_1c/2 receptor. The contractile effect was best mimicked by 5-MeOT suggesting a 5-HT₄-like receptor. The overall potency of agonists on the body wall muscle was 5-MeOT > 5-CT > 5-HT > α-Me-5-HT > 2-Me 5-HT > PAPP > Buspirone > > CGS-12066B.
• 4.4. Results suggest that either the sites of 5-HT action possess several different receptors or the receptors themselves are more complex with multiple properties.

     

Central serotonin level-dependent changes in body temperature following administration of tryptophan to pargyline- and harmaline-pretreated rats

• 1.1. The effect of tryptophan on body temperature was studied in rats pretreated with pargyline, an irreversible monoamine oxidase inhibitor (MAOI), and harmaline, a reversible MAOI.
• 2.2. Tryptophan (100 mg/kg IP) produced hypothermia followed by hyperthermia in pargyline-pretreated rats, and hypothermia in harmaline-pretreated rats, but tryptophan did not cause body temperature changes by itself.
• 3.3. The tryptophan-induced hypo- and hyperthermic effects, which peaked at about 1 and 6 hr after tryptophan administration, respectively, were accompanied by a significant increase in serotonin (5-HT) levels in the pargyline-pretreated rat brain (75%–138.7% and 207%–240.9% increase, respectively), and the 5-HT levels in the hyperthermic state were significantly higher than those in the hypothermic state.
• 4.4. In harmaline-pretreated rats, tryptophan also increased the central 5-HT levels (80.5%–95.5% increase) in the hypothermic state, and the effect peaked at about 1 hr after tryptophan administration. The central 5-HT levels in harmaline-pretreated rats slightly decreased at 6 hr after tryptophan administration and were significantly lower than those in the hyperthermic state in the pargyline-pretreated rats.
• 5.5. Tryptophan (100 mg/kg IP) administration decreased 5-hydroxy indole acetic acid (5-HIAA) levels, 5-HT turnover, and dopamine (DA) turnover in the brain of pargyline-pretreated rats, but these parameters were not significantly different between the hypothermic and hyperthermic states (i.e., at 1 and 6 hr after tryptophan administration, respectively).
• 6.6. These results suggest that the tryptophan-induced body temperature change depends on the different 5-HT levels in the brain and that the 5-HT level needed to induce hyperthermia is higher than that needed to induce hypothermia.

     

The reduction of antinociceptive effect of morphine administered intraventricularly is correlated with the decrease of serotonin release from the spinal cord in streptozotocin-induced diabetic rats

• 1.1. The antinociceptive effect of morphine (25 μg) administered into the 3rd ventricle was significantly attenuated in streptozotocin-induced diabetic rats as measured by the tail-flick assay.
• 2.2. The release of serotonin (5-HT; 5-hydroxytryptamine) from the spinal cord caused by intraventricular injection of morphine (25 μg) was significantly reduced in streptozotocin-induced diabetic rats.
• 3.3. No differences of 5-HT contents of the spinal cord (lumbar cord) between streptozotocin-induced diabetic- and vehicle-treated rats were found.
• 4.4. It is concluded that the reduction of antinociception produced by intraventricular injection of morphine in streptozotocin-induced diabetic rats might be, at least partly, due to the decrease of 5-HT release from the spinal cord.

     

Effects of μ-, δ- and κ-opioid antagonists in atrial preparations from nonfailing and failing human hearts

• 1.1. We examined the effects of naloxone (preferentially μ-antagonist), naltrindole (selective δ-antagonist) or nor-binaltorphimine (nor-BNI, selective κ-antagonist) on auricular myocardium tissue from nonfailing and failing human hearts.
• 2.2. The opioid antagonists used in this study induced inhibitory effects in auricular strips from failing and nonfailing human hearts. In addition, the maximal effect, the IC₅₀, and the slope of concentration-response curves obtained with μ-, δ-, and κ-opioid antagonists were similar in failing and nonfailing human heart tissues.
• 3.3. The K-antagonist was more effective than naltrindole or naloxone. Moreover, the IC₅₀ for nor-BNI (0.25±0.01 × 10⁻⁵M) was lower than the IC₅₀ for naloxone (26.5±5.0 × 10⁻⁵M) and naltrindole (13.8±2.0 × 10⁻⁵M). Similar results were obtained in auricular strips from failing human hearts.
• 4.4. Our results demonstrate that the failing heart does not modify the inhibitory cardiac effects obtained with selective opioid antagonists.