The effect of opioid agonists and antagonists on gastrointestinal motility in mice selected for high and low swim stress-induced analgesia.

BACKGROUND: The opioid system in the gastrointestinal (GI) tract plays an important physiological role, but is also responsible for the side effect of opioid drugs, including troublesome constipation in chronic pain treatment. The aim of this study was to characterize and validate a new mouse model to study the effects of opioid agonists and antagonists in the GI tract. METHODS: Six-week-old male Swiss-Webster mice, divergently bred for high (HA) and low (LA) swim stress-induced analgesia (SSIA), were used in the study. To assess the influence of opioid agonists (morphine and loperamide) and antagonists (naloxone hydrochloride, NLX and naloxone methiodide, NLXM) on GI motility, whole GI transit (whole GIT) and upper GIT assays were conducted. To evaluate the expression of opioid receptors in the ileum and colon of HA and LA mice, immune staining was performed. KEY RESULTS: The effect of morphine was more pronounced in HA line, whereas loperamide exerted a stronger effect in LA mice. Furthermore, NLX and NLXM differentially abolished the inhibitory action of the central and peripheral opioid system on whole and upper GIT in HA and LA mice. The differences in GI motility between HA and LA mice coexisted with parallel changes in the expression of opioid receptors in the ileum and colon. CONCLUSIONS & INFERENCES: Differences in the activity of the endogenous opioid system are responsible for the vulnerability to changes in GI motility during treatment with opioids. Our findings validate the HA/LA model for further studies on opioids in the GI tract.

Selective breeding of mice for high and low swim analgesia: differential effect on discrete forms of footshock analgesia.

Selective breeding of mice displaying high and low swim-induced analgesia led to the development of two animal lines divergent in the magnitude of analgesic response to swimming. In this study, animals belonging to the seventh generation of both lines were exposed to two temporally different forms of footshock, one of which produced opioid and the other non-opioid analgesia. We found that selective breeding for high and low swim-induced analgesia exerted a striking influence on the magnitude of the opioid-mediated type of footshock analgesia, but was ineffective on that of the non-opioid type.

Selective cone dystrophy with protan genotype.

PURPOSE: To determine the functional defects in two male patients with progressive cone dystrophy and hybrid L-M cone pigment genes. METHODS: Clinical evaluation, standard electroretinography, and electrooculography were performed in two affected patients and two family members. Measurements of spectral sensitivity and transient tritanopia were made in both patients. RESULTS: In the patients, visual acuity varied between 20/50 and 20/100. The electroretinogram showed reduced flicker responses. When light adapted, a-wave amplitudes were borderline, but b-wave amplitudes were reduced severely. Electroretinography with chromatic stimuli showed a difference between well-preserved responses to green and markedly reduced responses to red stimuli. Spectral sensitivity measurement revealed a lack of L (long-wavelength sensitive; red) cone function and normal function of the S (short-wavelength sensitive; blue) and M (middle-wavelength sensitive; green) cones. Transient tritanopia was abnormal, indicating a severe disturbance of cone-cone interaction. CONCLUSIONS: Progressive cone dystrophy with predominant dysfunction of L cones exists in both patients. The cone dystrophy may be caused by a rearrangement of the X-chromosome pigment gene array that is associated with the deletion of L-cone sequences and the formation of hybrid L-M cone pigment genes. It cannot be excluded, however, that both patients have protanopia and that cone dystrophy developed because of other causes.

Antidepressant-type effect of the NK3 tachykinin receptor agonist aminosenktide in mouse lines differing in endogenous opioid system activity.

The influence of the tachykinin NK3 receptor agonist, aminosenktide on the immobility in the forced swimming test was studied in mouse lines selectively bred for divergent magnitudes of stress-induced analgesia. The high analgesia (HA) line is known to display enhanced, and the low analgesia (LA) line displays reduced activity of the opioid system. Aminosenktide at doses of 125 microg/kg or 250 microg/kg intraperitoneally (IP) reduced, in naltrexone-reversible manner, the immobility more of opioid receptor-dense HA than of unselected mice, but was ineffective in the opioid receptor-deficient LA line. The effect of aminosenktide was quite similar to the antiimmobility action of desipramine (10 mg/kg IP), a prototypic antidepressant agent. None of the compounds increased animals' locomotion as found with an open field test; therefore their antiimmobility effect cannot be attributed to a change in general motility. The results claim that aminosenktide causes an antidepressant effect, and endogenous opioids are involved in this process.

Inheritance of stress-induced analgesia in mice. Selective breeding study.

Random mated Swiss mice swam for 3 min at 20 degrees C. The animals with less than or equal to 10 s and greater than or equal to 50 s postswim latencies on the hot plate (56 degrees C) were selected as progenitors of low (LA) and high (HA) analgesia lines, respectively. Gradual divergence of latency distributions and postswim hind paw flick latencies between the two lines was observed in successive generations. As shown by the tail flick test applied to the sixth offspring generation, postswim analgesia in the HA line was not only more pronounced but also lasted longer compared to the LA line and the unselected controls. Possible differentiation of the endorphin system activity in the course of selection is discussed.

Differentiation of neurochemical basis of stress-induced analgesia in mice by selective breeding.

Two selectively bred lines of mice, one responding with high (HA) and the other with low analgesia (LA) to 3 min swims at 20 degrees C were compared with respect to naloxone ability to reverse such swim induced pain threshold elevation. Also, the analgesic effect of morphine was tested in both lines of mice. Naloxone at a dose of 1 mg/kg significantly reduced postswim analgesia determined with hot plate and tail flick tests in HA mice, but was not effective in LA mice. HA mice responded with significant elevation of hot plate threshold to 0.1 mg/kg of morphine hydrochloride, whereas in LA mice comparable level of analgesia developed after 12.8 mg/kg. The data argue for greater involvement of opioid mechanisms in producing analgesia in HA mice than in LA mice, and so for inheritance of endorphin system(s) activity.

Levorphanol and swim stress-induced analgesia in selectively bred mice: evidence for genetic commonalities.

Two independent selective breeding programs have developed divergent lines of mice expressing either high and low swim stress-induced analgesia (HA/LA lines; Jastrzebiec, Poland) or high and low levorphanol analgesia (HAR/LAR lines; Portland, OR). In the present study, mice from both programs were tested for both levorphanol analgesia (2 mg/kg) and an opioid-mediated swim stress-induced analgesia (3 min swimming in 32 degrees C water) in the hot-plate test. Mice selected for high and low levorphanol analgesia displayed high and low swim stress-induced analgesia, respectively; mice selected for high and low swim stress-induced analgesia displayed high and low levorphanol analgesia, respectively. This pattern of correlated responses suggests a high degree of common genetic determination in opiate and swim stress-induced analgesia. These findings also suggest that individual differences in analgesic responsiveness to opiate drugs result from genetically determined individual differences in endogenous pain inhibitory mechanisms.

Mu and delta opioid receptor analgesia, binding density, and mRNA levels in mice selectively bred for high and low analgesia.

The present study examined mu and delta opioid analgesia, receptor binding, and receptor mRNA levels in lines of mice from two selective breeding projects of relevance to opioid analgesia. Large differences were observed in the analgesic potency of [d-Ala2, NMPhe4, Gly-ol]enkephalin (DAMGO), [d-Pen2,5]enkephalin (DPDPE), and [d-Ala2]deltorphin II (DELT), selective mu, delta1, and delta2 opioid receptor agonists, respectively, in mice selectively bred for high (HA) and low (LA) swim stress-induced analgesia (SIA). HAR and LAR mice, selectively bred for high and low levorphanol analgesia, respectively, display equally large differences in their analgesic sensitivity to DAMGO, modest differences in sensitivity to DPDPE, and no differences in sensitivity to DELT. These sizable genotypic differences in analgesic potency were accompanied by HA/LA and HAR/LAR differences in whole-brain homogenate [3H]DPDPE and/or [3H]DELT, but paradoxically not [3H]DAMGO, binding. Solution hybridization of mRNA extracts encoding mu (MOR-1) or delta (DOR-1) opioid receptors indicated some regional differences in gene expression between high and low lines. Surprisingly, differences in these in vitro markers were often in the direction of LAR>HAR. The present data indicate that selection for either SSIA or levorphanol analgesia produces differential effects on mu and delta opioid analgesia that are accompanied by alterations on in vitro assays, the significance of which remains to be determined. The data are discussed with regard to the utility of in vitro biological markers and genetic models of analgesia.

Mu-opiate receptor binding is up-regulated in mice selectively bred for high stress-induced analgesia.

Pain perception and sensitivity to opiate analgesics strongly depend on genotype. Mice selectively bred for high (HA) and low (LA) swim stress-induced analgesia display markedly divergent morphine analgesia, a difference that appears to be determined by one or at the most two major genes. In an attempt to provide candidate genes mediating the supranormal analgesia displayed by HA mice, we performed mu-opiate receptor binding on 27th generation HA, LA, and control (C) mice using [3H]naloxone. HA mice were found to have significantly higher whole-brain receptor density (Bmax) than LA mice in whole brain homogenates; no significant difference in affinity (Kd) was observed. Quantitative autoradiography confirmed the line difference in whole-brain receptor binding. In the medial thalamus, a brain area implicated in ascending pathways of pain inhibition, HA mice were found to display significantly higher [3H]naloxone binding than C mice (a 64% increase) and LA mice (a 128% increase). No significant line differences were observed in any other brain locus. Thalamic mu receptors may therefore play an important role in a central 'volume control' mechanism of pain inhibition, and underlie individual differences in the responses of mice to opiate analgesic drugs.

Antagonism of the non-opioid component of ethanol-induced analgesia by the NMDA receptor antagonist MK-801.

Recent evidence from our laboratory suggests that the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (dizocilpine) selectively antagonizes non-opioid (i.e. naloxone-insensitive) mechanisms of stress-induced analgesia in mice. For example, we have recently demonstrated that a low dose of MK-801 (0.075 mg/kg, i.p.) antagonizes the non-opioid component of a mixed opioid/non-opioid swim stress-induced analgesia (SSIA) resulting from forced swimming for 3 min in 20 degrees C water. Since ethanol-induced analgesia (EIA) has been found to be only partially attenuated by naloxone, we hypothesized that MK-801 would similarly block the non-opioid component of EIA. The effects of MK-801 and of the opioid receptor antagonist naloxone (10 mg/kg, i.p.) on analgesia produced by ethanol (2.5 g/kg in 20% vol/vol, i.p.) were studied in control mice and in mice selectively bred for high (HA) or low (LA) SSIA. HA mice showed significantly more, and LA mice significantly less, EIA than controls. Naloxone and MK-801 significantly attenuated EIA in control and HA mice, and in these lines the combined administration of both antagonists blocked EIA completely. In LA mice, which displayed very little EIA, naloxone but not MK-801 reversed EIA completely. These findings provide additional evidence for the role of the NMDA receptor in non-opioid mechanisms of analgesia. The finding that mice selectively bred for high and low SSIA, also display high and low EIA suggests common mediation of the effects of stress and ethanol on antinociceptive processes.

Morphine analgesia and tolerance in mice selectively bred for divergent swim stress-induced analgesia.

Morphine-induced analgesia and tolerance were examined in Swiss Webster mice selectively bred for high and low swim stress-induced analgesia. Morphine produced a dose-dependent analgesia in both lines; it was 4-fold more potent in the high analgesia line than in the low analgesia line. Despite the differences in morphine-induced analgesia, the degree of tolerance was the same in both lines. Together, these data suggest that selective breeding of mice for high and low swim stress-induced analgesia produced a striking difference in morphine-induced analgesia without affecting the degree of tolerance. Thus, while there is a common genetic determination in swim stress-induced and morphine-induced analgesia, the development of tolerance to morphine possibly relies on a different genetic background.

Acute morphine dependence in mice selectively-bred for high and low analgesia.

Acute morphine dependence was compared in mice selectively-bred for high (HA) and low (LA) swim stress-induced analgesia and high (HAR) and low (LAR) levorphanol analgesia by counting the number of naloxone-precipitated jumps. Whereas LAR mice displayed greater acute morphine dependence than HAR mice, HA and LA mice did not differ. No genotypic differences were observed in non-dependent mice, discounting possible differences in basal naloxone sensitivity and/or opioid peptide levels. Thus, the two selection projects, while both producing lines exhibiting highly divergent sensitivity to morphine analgesia, have not had analogous effects on all opioid measures, supporting the notion of independent genetic mediation of opioid analgesia and dependence. Further, these data suggest that analgesic sensitivity may not predict sensitivity to morphine dependence.

Cone and rod function in cone degenerations.

Progressive cone dystrophy (CD) is usually marked in the initial stages by reduced visual acuity, color vision deficiency and alterations in the photopic electroretinogram, while morphological alterations can be very mild; in some forms rods are affected in a later stage as well. We examined 40 patients with progressive cone dystrophy to determine the extent of functional losses in the cone system with psychophysical tests. A great variety of visual acuity and fundus alterations was found. Myopia was present in 74% of the patients. An autosomal dominant pattern of inheritance predominated (32%). No prevalence of gender was found. The age of onset ranged between 10 and 30 yr. All patients had progression of their symptoms. The total error score in color arrangement tests, the saturated Farnsworth Panel D-15 and the Farnsworth-Munsell 100-hue test, was pathologic with a predominance of confusions along the tritan and scotopic axis. Especially if visual acuity was below 0.5, color vision defects increased, but color vision defects were also found in patients with normal visual acuity. A general decrease of sensitivity in all three cone mechanisms was observed in measurements of spectral sensitivity. Moreover, cone-cone interaction as tested by transient tritanopia measurements was usually disturbed. In the dark adaptation function the threshold of the cone branch was usually elevated. These tests provide a good means to ascertain the correct diagnosis in early stages of the disease and to monitor progression in patients suffering from cone dystrophy.

Analgesia in selectively bred mice exposed to cold in helium/oxygen atmosphere.

In order to evaluate the stressing role of swim hypothermia in producing swim stress-induced analgesia (SSIA), we examined whether a mere decrease in the animals' core temperature without swimming would be sufficient to elicit analgesia. The subjects were Swiss-Webster mice selectively bred for 37 and 40 generations for divergent magnitudes of SSIA. High (HA) and low analgesia (LA) mice were exposed for 15 min to temperatures in the range between -5 and +20 degrees C in 79% He/21% O2 (Heliox) atmosphere. The Heliox exposure produced ambient temperature-dependent hypothermia and analgesia, as assessed with a hot-plate test (56 degrees C). The post-Heliox analgesia was of much higher magnitude in HA than in LA mice. The steeper slope of regression of the magnitude of analgesia upon hypothermia in HA mice indicates that these mice are far more sensitive to the analgesic effect of hypothermia than LA mice. Naltrexone HCl (10 mg/kg i.p.) attenuated analgesia in ambient temperature-dependent manner in HA, but not in LA mice. In view of the apparent similarity of Heliox-induced analgesia and SSIA we suggest that hypothermia is a powerful component of swim stress to induce SSIA in the mouse.

Adrenalectomy and dexamethasone differentially affect postswim antinociception in mice selectively bred for high and low stress-induced analgesia.

The effects of dexamethasone and naloxone on analgesia induced by swimming (3 min, 20 degrees C) were studied in the 6th and 7th generations of adrenalectomized and intact mice selectively bred for high (HA) and low (LA) postswim analgesia. Swim-induced analgesia in intact HA animals was significantly reduced by naloxone and dexamethasone while in LA mice these two compounds were ineffective. Naloxone ability to reverse adrenalectomy-caused swim analgesia increase was much greater in HA than in LA mice. In both intact and adrenalectomized HA animals dexamethasone and naloxone decreased postswim analgesia to the level observed in LA mice. It is suggested that selective breeding for high and low swim analgesia modified the extent of pituitary-adrenal axis involvement in the generation of stress-induced analgesia.

Acoustic startle and open-field behavior in mice bred for magnitude of swim analgesia.

Acoustic startle response (ASR) and open-field activity was examined in the 46th generation of mice that have been selectively bred for high analgesia (HA) and for low analgesia (LA) induced by 3-min swimming in 20 degrees C water. These lines were earlier found to differ in brain opioid receptor density and in the expression of opioid-mediated phenomena, as analgesic sensitivity to opiates and reversibility of swim stress-induced analgesia (SSIA) by naloxone. For comparison, a randomly bred control (C) line was used. To measure the amplitude of ASR, the mice were exposed to 110-dB acoustic stimuli in a Coulbourn apparatus. In saline-injected mice, the ASR force was found significantly lower in the LA than in the HA, as well in the C line, but did not differ between the two last lines. Naltrexone hydrochloride (10 mg/kg IP 30 min before ASR testing) augmented the startle in the opioid receptor-dense HA line, but had no effect in the opioid receptor-deficient LA line, as well in the C line; therefore, the ASR magnitude in naltrexone-injected HA mice was significantly higher compared to the C line. HA mice displayed less activity in an open-field test; that is, they remained immobile longer in the center of the field, and thereafter performed less ambulation and less rearing against the wall compared to the LA line. Naltrexone failed to modify the open-field activity in any line. The results confirm that the pattern of ASR depends on the genetic makeup of the animals. The higher amplitude of ASR, taken together with the lower open-field activity of HA mice, can be interpreted in terms of higher anxiety level, compared to the LA line. It is suggested that the higher ASR in HA mice relies on a nonopioid mechanism, which is tonically inhibited by the opioid system.

Opioid and nonopioid swim stress-induced analgesia: a parametric analysis in mice.

Environmental stress causes the activation of two types of endogenous pain inhibitory systems in animals: opioid analgesia is antagonized by opiate receptor blockers (e.g., naloxone and naltrexone), whereas analgesia produced by nonopioid systems is insensitive to such antagonism. A large literature documents that the parameters of the laboratory stressor will determine the neurochemical identity of the resultant analgesia. In rats, low severity stressors produce opioid analgesia and higher severity stressors produce nonopioid analgesia. A recent parametric analysis of swim stress-induced analgesia (SSIA) in the female Quackenbush mouse, however, observed the opposite pattern. The present study is a parametric analysis of SSIA using a range of swim temperatures (15-38 degrees C), swim durations (45 s to 7 min), and genetic models [male Swiss-Webster mice, and mice selectively bred from this outbred strain for high (HA), low (LA), or control SSIA]. We find that in nonselected mice low severity swims (i.e., warm temperature, short duration) produce naloxone-sensitive opioid SSIA, whereas high severity swims (i.e., cold temperature, long duration) produce nonopioid SSIA. This pattern is also seen in HA mice displaying very high analgesic magnitudes, but not in LA mice displaying minimal SSIA. In the selectively bred mice, analgesia and hypothermia from forced swimming are positively correlated, but can be dissociated both genetically and neurochemically. Furthermore, swimming in body temperature (38 degrees C) water produces analgesia without concommitant hypothermia, and the increased magnitude of 38 degrees C SSIA displayed by HA mice over control levels is entirely opioid.

Cross-tolerance between morphine and swim analgesia in mice selectively bred for high and low stress-induced analgesia.

Mice selectively bred for high (HA) and for low analgesia (LA) induced by 3-min swimming at 20 degrees C and unselected controls (C) were injected three times daily for 3 days with 20 mg/kg morphine HCl. The analgesic effect of 10 mg/kg morphine in nontolerant mice differed between the lines in the rank order of HA > C > LA and significantly decreased after repeated treatment with morphine, as revealed by the hotplate test (56 degrees C). The tolerance to morphine analgesia was more pronounced in HA than in C mice but did not develop at all in LA mice. Similarly, the magnitude of swim-induced analgesia in morphine tolerant mice decreased to a greater degree in the HA than the C line but did not change in LA mice. Naloxone HCl (1 and 10 mg/kg) attenuated swim analgesia more in nontolerant HA than C mice but had no effect in morphine-tolerant HA and C and in all LA mice. The differential degree of morphine tolerance and cross-tolerance with swim analgesia suggests that the strategy of selective breeding toward divergent magnitudes of stress-induced analgesia has differentiated opioid involvement in endogenous pain inhibition in the selected lines.

Potentiation of swim analgesia by D-amino acids in mice is genotype dependent.

The effect of combined treatment with 125 mg/kg of D-phenylalanine plus 125 mg/kg of D-leucine (IP) on magnitude and duration of analgesia caused by 3 min swim at 20 degrees C was studied in mouse lines selectively bred for 20 generations toward high and low level of stress-induced analgesia. The D-amino acids administered 30 min prior to swimming increased postswim tail-flick latencies and prolonged antinociception more in the high analgesia line (HA) than in concomitantly bred unselected controls, but were not effective in the low analgesia line (LA). The potentiation of swim analgesia by D-amino acids was prevented by simultaneous administration of 1 mg/kg of naloxone hydrochloride which, given alone, antagonized the analgesia more in the HA line than in controls, but not in the LA line. The results are interpreted in terms of genetic differentiation of opioidergic transmission in the selectively bred mouse lines.

Tolerance and cross-tolerance with morphine in mice selectively bred for high and low stress-induced analgesia.

Mice selectively bred for high (HA) and low (LA) swim-induced analgesia were exposed to two different stress paradigms; one consisting of a 3-min swim at 20 degrees C daily for 14 days, and the other consisting of 3-min swims repeated at 2-h intervals for 48 h. Both forms of chronic stress resulted in the development of tolerance to swim-induced antinociception to a greater degree in the HA mice than in control (C) mice, but were both ineffective at inducing tolerance in LA mice. Swimming repeated at 2-h intervals for 48 h resulted in cross-tolerance with morphine in HA and C mice. Naloxone (1 and 10 mg/kg, IP) failed to antagonize swim-induced analgesia in mice that had experienced chronic swimming in the 2-h/48-h paradigm. The daily swimming paradigm failed to produce cross-tolerance with morphine analgesia in any line. Differential degree of tolerance in three lines supports a hypothesis that selective breeding for high and low stress-induced analgesia has modified the degree of opioid involvement in the endogenous analgesia mechanisms.