Lab Mice Die After Drugs, Disco

[Kevster]

Linkage (http://story.news.yahoo.com/news?tmpl=story2&cid=518&u=/ap/20020819/ap_on_re_eu/britain_mice_experiment_3&printer=1)

Lab Mice Die After Drugs, Disco
Mon Aug 19, 7:49 AM ET

LONDON (AP) - The government on Monday reprimanded scientists who plied mice with drugs and loud dance music to study the effect on their brains.

The Home Office said it was taking "infringement action" against Cambridge University researchers who injected mice with the stimulant methamphetamine and subjected them to loud music, including tracks by dance act The Prodigy.

Several mice died and others suffered brain damage in the experiment, whose results were published in the journal NeuroReport last year.

Animal rights activists condemned the experiment. The British Union for the Abolition of Vivisection called it "tasteless and horrific."

The experiment was part of a wider study looking at the effect of amphetamine on a the striatum, a brain region that degenerates in Huntington's disease, a fatal, inherited brain disorder.

The findings suggested that loud pulsating noise like that found in dance clubs could intensify the drug's toxic effects.

Researchers studied 238 mice, injecting half with salt and half with the drug. While the mice injected with salt fell asleep when music was played, the drugged mice appeared to jiggle backward and forward.

Scientists found that the drugged mice suffered more speed-induced brain damage than normal. Seven mice who listened to the Prodigy died, as did four who were played music of a similar tempo by Bach.

The Home Office, the government department responsible for overseeing rules for animal research, did not say what form of action had been taken against the scientists.
______________________________________________________________________

I guess this might explain a few things about how Disco "died"...

[DoPeY5007]

:hmm: that is a very stange study :hmm:






and they injected them with salt :confused: wouldn't that kill ya?

[molecularfire]

Nope. I'm assuming by salt they meant saline. It's the equivalent of injecting them with water. Cells need a little salt in their water to do well. If you inject them with water, osmotic effects would draw the water into the cell making it swell and burst. Actually I don't see what is wrong with this experiment. I mean I haven't read the paper so I could be wrong, but heck, they did the proper controls and they got a finding that seems significant. What's wrong with what they did? :shrug:

[Napoleon54]

As Molecularfire said, this appears to be a valid, worthwhile study. Tons and tons of animal research is conducted every day. Animal studies are essential and can not be replaced. Unfortunately everytime a study involving animal research gets some publicity, the animal rights organizations throw a fit and portray it as being something obscene, out of the ordinary, and unnecessary. This could not be farther from the truth.

[Napoleon54]

Methamphetamine toxicity in mice is potentiated by exposure to loud music

A. Jennifer MortonCA; Miriam A. Hickey; Laura C. Dean

Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QJ, UK

NEUROREPORT 2001;12:3277-3281


--------------------------------------------------------------------------------

Methamphetamine (METH) is a drug of abuse used for its stimulant effects. Its neurotoxicity is very variable, and is increased by a number of factors, including crowded conditions and increased ambient temperature. The effects of such factors are increasingly important, with the widespread use of these stimulants at nightclubs and 'raves'. Here, we compared the effect of another dominant feature of nightclubs, continuous loud noise, on the toxicity of METH in mice. We found that mice exposed to loud music exhibited longer lasting stereotypy, an altered place preference in the open field and had more seizures than mice given METH in a quiet setting or when exposed to loud white noise. A greater increase in reactive gliosis was also seen after exposure to METH and loud music. Thus, METH appears to be more toxic when taken while exposed to loud music.

Key words: Behaviour; Damage; Gliosis; Methamphetamine; Mice; Music; Striatum



--------------------------------------------------------------------------------


INTRODUCTION
The toxic potential of METH and its derivatives has been known for many years. With METH already the third most popular stimulant drug used in the USA, and used with increased frequency at the club scene in Europe, the factors that potentiate METH toxicity are of considerable importance [1,2]. The effects of ambient temperature and aggregation on toxicity of METH and ecstasy have been well studied [2-6]. However, the potentiating effect of loud noise on METH toxicity was last investigated in the 1940s [7], and the noise used in these studies was a sudden, occasional noise, such as intermittent banging of a metal box. As well as being inexact and difficult to replicate, this kind of noise is unlikely to mimic the noise to which METH users are exposed at nightclubs or raves, where loud music with a strong percussive beat is played continuously. In this study therefore, we examined the effect of continuous loud music on the toxicity of METH in mice.

MATERIALS AND METHODS
Behavioral testing:

All experiments were carried out under the authority of a licence approved by the local ethics committee and issued by the Home Office. We used groups of 9 or 10 music-naive, adult C57/BL6 mice. Experiments were conducted at an ambient temperature of 22°C and were video-recorded for analysis by a blinded observer. Each experiment was repeated at least three times. Mice were placed in an open field, and, after a 30 min habituation period, were given METH (75 mg/kg s.c.). They were then returned to the open field for 3 h. Control mice received saline. We used four different noise conditions: (i) silence (ambient noise; 55 dB), (ii) loud white noise (95 dB) (iii) rave music (Prodigy, average loudness 95 dB) and (iv) classical music (Bach, average loudness 95 dB). Rave music consisted of four tracks from The Prodigy. Note that rave music was chosen for its musical qualities (as a popular music genre with a recognizable, percussive signature that is frequently played very loudly) rather than because of its association with raves. Classical music was the Allegro movement from J. S. Bach's Violin Concerto in A minor (BWV 1041). Music was edited to retain tempo and loudness and was repeated continuously. Noise intensity was measured using a Solex decibel monitor. The average loudness of 95 dB is similar to the maximum loudness of personal stereos but considerably lower than the 105-115 dB found in discos, nightclubs and rock concerts [8].

Analysis parameters measured were seizure activity [9], locomotion in the open field [10] and stereotypy [11]. Locomotion was determined by counting the number of squares each mouse entered at different times after METH administration. Data were pooled in 30 min bins, and expressed as means ± s.e.m. The significance of the data was determined using a one way ANOVA followed by Newman-Keuls post-hoc test. Significance of seizure activity and survival was tested using contingency tables; p values were calculated using Fisher's exact test. A critical value for significance of p < 0.05 was used throughout the study.

In the original noise experiments conducted by Chance [7], amphetamine was administered to single mice at high doses (150 or 180 mg/kg), and its toxicity was determined by the increase in mortality. We wanted to repeat these experiments using a subtoxic dose of METH. Thus, in a series of pilot studies, METH (12, 25, 50, 65, 75, 85, 100 and 125 mg/kg) was administered s.c. to groups of mice to determine an appropriate dose (data not shown). The dose used here (75 mg/kg) was chosen because it was the highest dose tested that did not kill any mice in the pilot experiments.

Immunocytochemistry:

Mice were killed 1, 3, 7, 14 or 35 days after the experiment, and serial sections of their brains were examined immunocytochemically for reactive astrogliosis using GFAP. Brain sections from kainic acid-treated mice (20 mg/kg, i.p., killed 1 week after injection) were used as controls for seizure-induced reactive gliosis. Cryosections of brains were stained for GFAP, and visualized using diaminobenzidine. GFAP-positive astrocytes were counted in striatum on both sides of the brain in 12 fields (four from each of three sections) from 3-5 animals in each group.

RESULTS
Locomotor activity in the open field:

Mice treated with saline were not affected by the loudness of the noise or the type of noise stimulus (Fig. 1a, upper panels). Indeed, within 30 min of the saline injection, most of the mice were asleep, and remained so for the rest of the experiment. In contrast, we found that METH-induced behavior of the mice varied markedly with different noise stimuli (Fig. 1a, lower panels). When given METH in silence or with white noise, mice exhibited behavior typical of METH intoxication [12-15] with an initial period of hyperactivity followed by stereotypic behavior resulting in an altered place preference and reduced locomotion. Stereotypy lasted for 60-90 min, after which the mice returned to a hyperactive state. In contrast, mice exposed to METH/loud music rapidly became stereotypic and remained so for 2 h. There was significantly less locomotion with both types of music compared to white noise from 120 min onwards (p < 0.001). As well as being longer lasting, the METH/music-induced stereotypy was different from that seen with silence/white noise (Fig. 1b). In particular, METH/music caused backward walking and circling that was not seen in the METH/silence or METH/white noise groups. The behavior was similar in mice from both music groups, although those in the Prodigy group moved in tighter circles. Some circling was also seen with mice given METH in silence or white noise. However the circling in these mice was qualitatively different, in that the mice rarely completed multiple 360° turns, and moved forwards as well as backwards.


--------------------------------------------------------------------------------
Fig. 1. The effect of saline (upper panels) or METH (lower panels) on locomotion in the open field (a). The solid bar shows the duration of the sterotypic behavior. (b) Patterns of activity of two typical mice (red and blue lines) were traced for a 5 min period starting 10 min, 60 min or 2 h after METH (or saline) was administered.


--------------------------------------------------------------------------------

The change in stereotypy seen here suggests that exposure to loud music alters the balance of neurotransmitters released after METH. METH produces its stimulatory effect by promoting the release of biogenic amine transmitters [16], in particular dopamine, but also 5-hydroxytryptamine (5-HT) and noradrenaline. METH-induced stereotypy has been well characterized [12-15,17] and it is believed that DA and 5-HT mediate different aspects of stereotyped behavior. DA-dependent behaviors include forward locomotion and head bobbing (seen with METH/silence or white noise), while 5-HT-dependent behaviors include forepaw treading, wet dog shakes and Straub tail (only occasionally seen in our mice). However, backward walking (seen with METH/music) is thought to require activation of both DA and 5-HT release. Thus our data suggest that music causes a change in METH-induced stereotypy from a predominantly DA-mediated stereotypy to one mediated by both 5-HT and DA.

Place preference in the open field:

Other behavioral parameters were differentially affected by different noise conditions. In particular, mice exposed to METH and music showed altered place preference compared with mice given METH in silence or white noise, with mice exposed to Prodigy and Bach spending proportionally more time in the inside squares than mice in any of the other groups (Fig. 2). This was most pronounced in the Prodigy mice. By 30 min these mice spent significantly more time in the center squares than mice exposed to other kinds of noise (p < 0.001 for silence, p < 0.01 for Bach and white noise). By 60 min mice from all groups spent significantly more time in the center squares, and there was no difference in center preference between groups (p > 0.05 for all). However by 90 min, while the center preference of the Prodigy group remained high, that of the silence and white noise groups had returned to baseline. By 120 min the center preference of the Bach group was also significantly lower than the Prodigy group (p < 0.05) and not significantly different from the silence or white noise groups (p > 0.05). Altered place preference in the open field has been related to altered levels of anxiety [18], with an increased preference for the inside squares used as measure of decreased anxiety. Note however that while these changes in behavior are consistent with an increase in 5-HT release, mice exposed to music also became stereotypic more rapidly than mice given METH in silence or white noise. Thus, their level of awareness and thus of place may be blunted by the METH-induced stereotypy.


--------------------------------------------------------------------------------
Fig. 2. The effect of METH/noise on center preference in the open field. There was no difference between center preference of any group in the 30 min before METH was administered. After administration of METH, center preference was significantly altered. ***p < 0.001, **p < 0.01, *p < 0.05 compared to silence. Where no error bars are shown, they are within the symbols.


--------------------------------------------------------------------------------

[Napoleon54]

Seizures and survival:

METH can be neurotoxic [2,3,19-22], and intermittent loud noise has been reported to increase this toxicity [7]. At high doses, METH causes seizures, and we used seizure activity [9] as a behavioral index of toxicity. Some mice from all METH-treated groups exhibited seizures. However the toxicity of METH was potentiated in mice exposed to loud noise, since mice given METH while exposed to white noise, Prodigy or Bach exhibited more seizures than mice given METH in silence (p < 0.05 white noise; p < 0.001 Prodigy and Bach; Table 1). There was no difference between the number of seizures experienced by mice in the Prodigy and Bach groups. There was also an increased mortality associated with METH/music, but not METH/silence or white noise (p < 0.05; Table 1). Together, these data suggest that exposure to continuous loud music increases the toxicity of METH.


--------------------------------------------------------------------------------
Table 1. Effect of noise on METH-induced seizures and mortality.


--------------------------------------------------------------------------------

Immunohistopathology:

After the experiment, there were no obvious long-lasting effects of METH/loud noise on mouse behavior in the home cage. Nevertheless, we wanted to see if there were any long-term effects of METH/noise exposure on the brain. We examined the brains of all mice for reactive gliosis, a marker of neurological damage. An increase in GFAP-positive staining was seen in the striatum of all mice given METH (Fig. 3). This is consistent with findings of others showing that METH increases striatal gliosis [4,21]. The increase in gliosis was independent of whether or not the mouse exhibited seizures, and indeed, the pattern of GFAP immunoreactivity contrasts with that seen in mice that experienced kainic acid-induced seizures, where striatal reactive gliosis was minimal. However, the time-course for astrocytosis differed according to the noise parameter. In METH/silence mice, GFAP immunoreactivity was maximal at 3 days, significantly reduced by 7 days (p < 0.05) and had disappeared completely by 2 weeks. In contrast, the glial response in METH-treated mice exposed to loud white noise and loud music was still elevated 7 days after the experiment. After 2 weeks, astrocytosis in brains of mice exposed to white noise had returned to background levels, although it remained elevated in the METH/music brains. Indeed, 5 weeks after the experiment was performed, a persistent striatal gliosis was present in the brains of METH/Prodigy-treated mice (48 ± 15 astrocytes/mm2, n = 3). (Astrocytosis in METH/Bach brains was elevated at 2 weeks, but was not examined at 5 weeks). It should be noted also that six of the music-treated mice died within 24 h of the experiment, and their brains were not included in the histological study. Therefore, if anything, the toxic effects of the combination of METH/music on reactive gliosis are likely to be under-estimated.


--------------------------------------------------------------------------------
Fig. 3. Photomicrographs (a) showing reactive gliosis in sections of brains of mice killed 7 days after exposure to METH. The increased GFAP-positive immunoreactivity in the striatum is shown at higher magnification in the right hand panels. Note that the morphology of astrocytes in striatum exposed to noise and music was different from that of astrocytes in striatum of METH/silence or kainic acid (KA)-treated mice, with stronger GFAP immunoreactivity and conspicuous processes. Bar = 25 m. Quantification of gliosis (b) showed a significantly higher number of GFAP-positive astrocytes in the striatae of mice killed 3 or 7 days after receiving METH compared to saline-treated mice. S = silence, WN = white noise, P = Prodigy, B = Bach. **p < 0.05 compared to WN, P or B.


--------------------------------------------------------------------------------

The fact that METH-induced toxicity was potentiated more by loud music than by loud white noise suggests that it is the music, rather than the loudness per se, that causes the effect. It could be argued that the mice habituated to white noise but not to the music, and that we were simply observing a stress-induced response [10] in the music groups. However, this seems unlikely, since in the saline-treated groups, all mice from all groups spent most of the experiment asleep, suggesting that they habituated as easily to loud music as they did to white noise. Further, none of the saline treated mice died or had seizures, and no reactive gliosis was seen in the striatae of mice from any saline-treated group. Thus prolonged exposure to loud music augments the neurotoxic effect of METH, but does not have a direct neurotoxic effect of its own.

The mice used in this study were music-naive, and it seems likely that they perceived both kinds of music simply as percussive noise, rather than as a particular genre of music. However, the clinical relevance of our findings is likely to be much greater for popular music styles than for classical music. Classical music is rarely played very loudly in public places and there is no evidence of an association between listening to classical music and stimulant abuse. In contrast, there is a relationship between drug use and music style, with fans of rave music being more likely to have experimented with drugs than those who preferred other styles of music [23]. There is also a relationship between the behavioral characteristics of people who listen excessively to loud music and those who abuse substances [24].

CONCLUSION
Our study shows that exposure not only to loud noise, but loud music in particular, potentiates the toxicity of METH. This has clear implications for the use of METH in noisy settings. However, there are caveats that must be applied before our findings are extrapolated to humans. First, our mice had no option but to be exposed to loud music. Thus, the circumstances under which they were given METH were likely to be more stressful than those of humans who go to clubs and voluntarily expose themselves to loud music. Second, while there is considerable evidence for the toxicity of amphetamines and its derivatives in rodents [21] and brains of human addicts [25], the long-term effects on the human brain of occasional recreational use of METH are uncertain. The mechanisms underlying the METH/music-induced changes in mouse behavior and brain pathology seen here, are also not known. Finally, our data should not be extrapolated to encompass other party drugs such as ecstasy, although clearly in view of our findings, a detailed study of the toxicity of ecstasy/loud music would be very interesting.

Acknowledgements:
L.C.D. was supported by a GIAP Summer studentship. We thank W. Leavens for histological assistance and R. Hart for photographic work.

REFERENCES


Pulse Check. Trends in Drug Abuse, Mid-Year Executive Office of the President. Office of National Drug Control Policy. Published online at www.whitehousedrugpolicy.gov
Green AR, Cross AJ, Goodwin GM. Psychopharmacology (Berl) (1995) 119, 247 -260.
Chance MRA. J Pharmacol Exp Ther (1946) 87, 214 -219.
Fink GB, Larson RE. J Pharmacol Exp Ther (1962) 137, 361 -364.
Craig AL, Kupferberg HJ. J Pharmacol Exp Ther (1972) 180, 616 -624.
O'Callaghan JP, Miller DB. J Pharmacol Exp Ther (1994) 270, 741 -751.
Chance MRA. J. Pharmacol Exp Ther (1947) 89, 289 -296.
Liebel J, Delb W, Andes C, Koch A. Laryngo-Rhino-Otologie (1996) 75, 259 -264.
Racine RJ. Electroencephalogr Clin Neurophysiol (1972) 32, 281 -294.
MacLennan AJ, Maier SF. Science (1983) 219, 1091 -1093.
Costall B, Naylor RJ, Olley JE. Eur J Pharmacol (1972) 18, 83 -94.
Fog R. Acta Neurol Scand Suppl (1972) 50, 3 -66.
Kuczenski R, Segal SD, Aizenstein ML. J Neurosci (1991) 11, 2703 -2712.
Curzon G, Femando JCR, Lees AJ. Br J Pharmacol (1979) 66, 573 -579.
Andrews CD, Fernando JCR, Curzon G. Neuropharmacol (1982) 21, 63 -68.
Sulzer D, Chen TK, Lau YY. et al. J Neurosci (1995) 15, 4102 -4108.
Sonsalla PK, Giovanni A, Sieber BA. et al. Ann NY Acad Sci (1992) 648, 229 -238.
Crawley JN. Neurosci Biobehav Rev (1985) 9, 37 -44.
Jonsson G, Nwanze E. Br J Pharmacol (1982) 77, 335 -345.
Seiden LS, Sabol KE. NIDA Res Monogr (1996) 163, 251 -276.
Wrona MZ, Yang Z, Zhang F, Dryhurst G. NIDA Res Monogr (1997) 173, 146 -174.
Fumagalli F, Gainetdinov RR, Valenzano KJ. et al. J Neurosci (1998) 18, 4861 -4869.
Forsyth AJ, Bamard M, McKeganey NP. Addiction (1997) 92, 1317 -1325.
Florentine M, Hunter W, Robinson M. et al. Ear Hear (1998) 19, 426 -428.
Volkow ND, Chang L, Wang G-J. et al. Am J Psychiatry (2001) 158, 377 -382.
CACorresponding Author

Received 1 August 2001;

accepted 9 August 2001

[Napoleon54]

http://ipsapp003.lwwonline.com/journals/neuroreport2/issues/2001-12-15/texts/f20011029002301.jpg
Fig. 1. The effect of saline (upper panels) or METH (lower panels) on locomotion in the open field (a). The solid bar shows the duration of the sterotypic behavior. (b) Patterns of activity of two typical mice (red and blue lines) were traced for a 5 min period starting 10 min, 60 min or 2 h after METH (or saline) was administered.

http://images.snapfish.com/334%3C%3A48323232%7Ffp65%3Dot%3E2326%3D879%3D86%3C%3Dxroqdf%3E23232%3C%3A%3B%3A66%3B9ot1lsi
Fig. 2. The effect of METH/noise on center preference in the open field. There was no difference between center preference of any group in the 30 min before METH was administered. After administration of METH, center preference was significantly altered. ***p < 0.001, **p < 0.01, *p < 0.05 compared to silence. Where no error bars are shown, they are within the symbols.

http://images.snapfish.com/334%3C%3A48323232%7Ffp66%3Dot%3E2326%3D879%3D86%3C%3Dxroqdf%3E23232%3C%3A%3B%3A66%3C%3Bot1lsi
Table 1. Effect of noise on METH-induced seizures and mortality.

http://images.snapfish.com/334%3C%3A48323232%7Ffp67%3Dot%3E2326%3D879%3D86%3C%3Dxroqdf%3E23232%3C%3A%3B%3A66%3C%3Aot1lsi
Fig. 3. Photomicrographs (a) showing reactive gliosis in sections of brains of mice killed 7 days after exposure to METH. The increased GFAP-positive immunoreactivity in the striatum is shown at higher magnification in the right hand panels. Note that the morphology of astrocytes in striatum exposed to noise and music was different from that of astrocytes in striatum of METH/silence or kainic acid (KA)-treated mice, with stronger GFAP immunoreactivity and conspicuous processes. Bar = 25 m. Quantification of gliosis (b) showed a significantly higher number of GFAP-positive astrocytes in the striatae of mice killed 3 or 7 days after receiving METH compared to saline-treated mice. S = silence, WN = white noise, P = Prodigy, B = Bach. **p < 0.05 compared to WN, P or B.

[kimchicowboy]

i saw some of the monkeys and cats research is done on at UCLA. pretty sad sight. but i guess all in the name of science, eh?

[hapoo]

humans do it anyway, why not study them?

[Napoleon54]

Originally posted by hapoo
humans do it anyway, why not study them?

For starters, it's illegal to give humans methamphetamine.


Oh yeah, and human subjects prolly wouldn't like for you to cut open their skulls and pull out their brains afterwards. :)

[Merlin]

Originally posted by Kevster
...The Prodigy....

See, this is just another way for tha MAN to try and keep one of the greatest bands out there down.


SMACK MY BITCH UP.

[molecularfire]

Originally posted by hapoo
humans do it anyway, why not study them?

I asked that once when I was working in a cancer lab. I mean... let's face it... the stuff we learn will never go towards helping all of mousekind. They just gave me a funny look and took away my scalpel. :(