Učinak pentadekapeptida BPC 157 na senzorno-motorni oporavak štakora nakon primjene intravenskih anestetika [The antagonistic effect(s) of pentadecapeptide BPC 157 in general anaesthesia in rats]

Zemba, Mladen (2008) Učinak pentadekapeptida BPC 157 na senzorno-motorni oporavak štakora nakon primjene intravenskih anestetika [The antagonistic effect(s) of pentadecapeptide BPC 157 in general anaesthesia in rats]. PhD thesis, Sveučilište u Zagrebu.

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Abstract

The aim of this investigation was to inquire the antagonism activity of pentadecapeptide BPC 157 on general anaesthetic activity of diazepam, propofol, thiopental, ketamine, fentanyl, dehidrobenzperidol and thalamonal in rats, and if observed, should enable information about mechanism of action. Pentadecapeptide BPC 157 was applicated firstly alone. Pentadecapeptide BPC 157 (10 pg kg-1, or 10 ng kg-1 or 10 µg kg-1) or the equal volume of physiologic solution were applicated 5 min before the various anaesthetics (kg-1 b. w.): diazepam (10.0; 15.0; 20.0 mg), propofol (50.0; 75.0; 100.0 mg), thiopental (20.0; 30.0; 40.0 mg), ketamine (50.0; 75.0; 100.0 mg), fentanyl (0.20; 0.30; 0.40 mg), dehydrobenzperidol (6.0; 8.0 mg) and thalamonal (0.1 + 5.0; 0.15 + 7.5 mg). All the substances or physiologic solution were given in acute application, in volume 5 mL/kg intraperitoneally (i.p.). In the experiments were used 6 rats per group, total 912 rats. All rats were used only once in the experiment. The rats were examined in the cage and in the open field. General anaesthetic drugs activity was determined according to the loss of righting reflex assessed in 5 min. intervals, as the indicator of anaesthesia. Mean anaesthetic scores (MASs) were calculated from mean of scores recorded every 5 minutes after anaesthetic injection to recovery the loss of righting reflex in all animals in group for a maximum of 2 h. The time required to return to a normal righting reflex was considered the recovery time (min). The number of animals losing the righting reflex of the total that received a specific treatment was used to calculate the percentage of loss of the righting reflex. ED50 for loss of the righting reflex, 95% confidence limits of that value, and the significance of differencies in ED50 values between groups were determined according to the method of Litchfield and Wilcoxon. Somatosensory (dis)orientation was evaluated in separate groups in rats according to their reaction to the standardized compression of the tail root in 10 min intervals. Responses were assessed using simple scoring system, described by Jelovac et al., (1998). Motor and locomotor behavioral responses were assessed in 10 min intervals in a cage and open field using simple scoring system, described by Ellinwood and Balster (1974). The cataleptic status of rats was assessed by placing animals in a rearing position. If the rats maintained cataleptic posture for more than 20 s, it was scored as cataleptic. The time during which the rats maintained this immobile position was recorded cumulatively up to 120 min. By application of BPC 157 alone, we did not note any change in behavior of the rats. BPC 157 antagonized sedative/hypnotic effects of diazepam. ED50 was significantly increased from 15.1 (13.7-16.5) to 17.3 (15.8-18.9) mg kg-1 (p<0.05). The changes in the ED50 values for diazepam were + 14 %, after all doses of BPC 157 respectively. The MASs, the number of rats with loss of righting reflex, and the recovery time were decreased (p<0.05). Relative change in recovery time for diazepam was 64%. Sedative/hypnotic effect of diazepam were decreased (p<0.05), and activity in place was increased (p<0.05). Propofol increased the rate of rats with loss of righting reflex in a dose-dependent fashion with ED50 82.6 (73.6-92.6). General anaesthetic activity of propofol was not changed with BPC 157. In ng-µg BPC group by propofol 100 mg kg-1 somatosensory orientation was improved 5 min and motor activity 40 min after propofol administration (p<0.05). General anaesthetic potency calculated by ED50 value for thiopental, was significantly decreased for 10 ng kg-1 from 21.1 (10.1-32.1) to 39.5 (26.2-67.8) (p<0.05) and for 10 μg kg-1 from 21.1 (10.1-32.1) to 37.2 (24.3-62.3) (p=0.05). The changes in the ED50 values for tiopental were + 87 % after 10 ng kg-1 and + 76 % after 10 μg kg-1 of BPC 157 respectively. MASs, the percentage of rats with loss of righting reflex, and recovery time were significantly decreased (p<0.05). Relative change in recovery time for thiopental was 89 %. Somatosensory orientation was improved after 50 min, and motoric recovery was significantly improved by ng-µg doses of BPC 157 after 70-80 min (p<0.05 and p<0.001). Ketamine increased the rate of rats with loss of righting reflex in a dose-dependent fashion with ED50 73.6 (65.2-82.5). General anaesthetic activity of ketamine was not changed with BPC 157. BPC 157 generally improved somatosensory dysorientation of ketamine 50 min after injection (p<0.05 and p<0.001). BPC 157 improved and changed the motoric recovery 70-80 min after injection (p<0.001 and p<0.05). Fentanyl induced a cataleptic response by fentanyl 300 µg kg-1. Catalepsy started about 1-2 min after fentanyl injection and lasted 22,23 ± 3,56 min. All rats displayed the Straub tail reaction with prominent muscular rigidity. The rats did not show catalepsy after fentanyl 200 µg kg-1. Rats given fentanyl 400 µg kg-1 displayed involuntary body torsion with hind paws in abduction, and lethal outcome. BPC 157 significantly (p<0.05) shortened catalepsy induced by fentanyl 300 µg kg-1 to 5.12 ± 1.29 and 4.15 ± 1.26 min. Relative change in recovery from fentanyl catalepsy was 81%. In difference, BPC 157 did not have any positive impact on recovery of rats administered fentanyl 400 µg kg-1. Dehydrobenzperidol at doses 6 and 8 mg kg-1 induced dose-dependent cataleptic action. Dehydrobenzperidol 6 mg kg-1 produced a cataleptic response with duration of 60,45 ± 2,25, and was about 3-fold greater than that produced by fentanyl 300 µg kg-1. BPC 157 significantly decreased catalepsy in rats treated by dehydrobenzperidol (6.0 and 8.0 mg kg-1) (p<0.05). Relative change in recovery time for droperidol was 100 %. Treatment with pentadecapeptide BPC 157 decreased catalepsy by lower dose (fentanyl 0,1 mg kg-1 + droperidol 5,0 mg kg-1) of thalamonal from 20,00 ± 2,28 min on 0,00 ± 0,00 min. Pentadecapeptide BPC 157 has no effect on the normal animals, but it is effective in animals with disturbations caused by i. v. general anaesthetics. The activity of CNS, significantly changed by effects of BPC 157, caused the change of anaesthetic potency of thiopental, and the cange of hypnotic potency of diazepam. BPC 157 also decreased the catalepsy time caused by fentanyl, droperidol and thalamonal. But, it not changed the anaesthetic potency of propofol and ketamine. BPC 157 corrects the disturbances which appear by anesthetics, in whose activity is the NO/cGMP transmission paticular important, and it interfere also biological impact of other neurotransmitter systems, whose activity is linked with its primary effects.

Abstract in Croatian

U ovom istraživanju željelo se ispitati antagonizam pentadekapeptida BPC 157 na opće anestetsko djelovanje diazepama, propofola, tiopentala, ketamina, fentanila, droperidola i talamonala u štakora. Pentadekapeptid BPC 157 je prvo apliciran sam u dozi 10 μg/kg i. p. Pentadekapeptid BPC 157 (10 pg, ili 10 ng ili 10 μg/kg t.t.) ili jednak volumen fiziološke otopine davani su 5 minuta prije različitih anestetika (/kg t.t.): diazepama (10,0; 15,0; 20,0 mg), propofola (40,0; 75,0; 100,0 mg ), tiopentala (20,0; 30,0; 40,0 mg), ketamina (50,0; 75,0; 100,0 mg), fentanila (200,0; 300,0; 400,0 µg), dehidrobenzperidola (6,0; 8,0 mg), talamonala (kombinacije fentanila i dehidrobenzperidola) (0,1 + 5,0 mg; 0,15 + 7,5 mg). Sve tvari ili fiziološka otopina davani su u akutnoj primjeni, u volumenu 5 mL/kg aplicirani intraperitonealno (i. p.). U pokusima se koristilo 6 ženki Wistar štakora težine 200-250 g po skupini, ukupno 912 štakora. Štakori su ispitivani individualno u kavezu i slobodnom polju. Svi štakori su korišteni samo jedanput u pokusu. Opća anestetska / anesteticima slična potencija lijekova ispitivana je koristeći gubitak refleksa uspravljanja kao indeks anestezije. Prosječna ocjena anestezije ili anesteziji sličnog učinka (POA) računala se iz prosjeka ocjena bilježenih svakih 5 minuta nakon injekcije anestetika do oporavka refleksa uspravljanja u svih životinja u skupini do najviše 2 sata. Vrijeme potrebno za povratak normalnog refleksa uspravljanja smatrano je vremenom oporavka (min). Broj štakora koji su izgubili refleks uspravljanja od ukupnog broja koji su dobili specifični tretman korišten je za izračunavanje postotka gubitka refleksa uspravljanja. ED50 gubitka refleksa uspravljanja, interval povjerenja 95%-ne pouzdanosti i pouzdanost razlika u ED50 vrijednostima među grupama određivani su po metodi Litchfield-a i Wilcoxon-a. Učinak anestetika na pokretljivost i somatosenzornu orijentaciju procjenjivan u 10 min intervalima, na zasebnim grupama štakora, prema reakciji na standardiziranu kompresiju repa. Odgovori su procjenjivani korištenjem jednostavnog bodovnog sustava Jelovca i sur., (1998). Motorni i lokomotorni odgovori ocijenjivani su u 10 min intervalima u kavezu i otvorenom polju korištenjem jednostavnog bodovnog sustava Ellinwood-a i Balster-a (1974). Vrijeme katalepsije određivalo se nakon i. p. primjene fentanila, dehidrobenzperidola i talamonala. Stanje katalepsije štakora procjenjivano je postavljanjem štakora u položaj uzdizanja postavljanjem prednjih šapa na ljestvicu. Ukoliko su štakori zadržavali kataleptički položaj duže od 20 sekundi, to je ocjenjivano katalepsijom. Vrijeme za koje su štakori održavali to imobilno stanje bilježilo se kumulativno do 120 min. Aplikacijom BPC 157 10 μg/kg i. p. samog nisu izazvane nikakve promjene. Primjenom BPC 157 i. p. antagonizirano je sedativno/hipnotsko djelovanje diazepama. ED50 za diazepam značajno je povećana sa 15,1 (13,7-16,5) na 17,3 (15,8-18,9) mg/kg u prisustvu pentadekapeptida BPC 157 (p<0,05). Promjena ED50 vrijednost za diazepam iznosila je + 14 % nakon sve tri doze pentadekapaptida BPC 157. Aplikacijom BPC 157 smanjena je POA i postotak štakora s gubitkom refleksa uspravljanja i 64 % skraćeno vrijeme oporavka (p<0,05) inducirano s diazepamom. Primjenom BPC 157 značajno je poboljšana somatosenzorna orijentacija u prvih 20 minuta (p<0,05) i aktivnost štakora u mjestu od 20-40 minute (p<0,05 i p<0,001). Propofolom je povećan postotak gubitka refleksa uspravljanja na način ovisan o dozi s vrijednošću ED50 82,6 (73,6-92,6). Primjenom BPC 157 nije promijenjeno opće anestetsko djelovanje propofola. Primjenom BPC 157 poboljšana je somatosenzorna orijentacija u 5 min (p<0,05), te od 40 minute i motorna aktivnost (p<0,05) kod propofola 100 mg / kg t. t. Primjenom BPC 157 signifikantno je antagoniziran anestetski učinak tiopentala. Djelovanjem tiopentala povećan je postotak gubitka refleksa uspravljanja na način ovisan o dozi s vrijednošću ED50 21,1 (10,1-32,1). Opća anestetska potencija kalkulirana preko ED50 vrijednosti za tiopental, značajno je smanjena za 10 ng / kg BPC 157 sa 21,1 (10,1-32,1) na 39,5 (26,2-67,8) (p <0,05) i za 10 μg / kg BPC 157 sa 21,1 (10,1-32,1) na 37,2 (24,3-62,3) (p=0,05). Promjena ED50 vrijednost za tiopental iznosila je + 87 % za 10 ng/kg i + 76 % za 10 μg/kg BPC 157. Uz BPC 157 značajno je smanjena POA i postotak štakora s gubitkom refleksa uspravljanja i 89 % skraćeno vrijeme oporavka (p<0,05). Somatosenzorna orijentacija i motorni oporavak značajno su poboljšani uz BPC 157 u cijelom periodu promatranja (p<0,05 i p<0,001). Djelovanjem ketamina povećan je postotak gubitka refleksa uspravljanja na način ovisan o dozi s vrijednošću ED50 73,6 (65,2-82,5). Opća anestetska potencija kalkulirana preko ED50 vrijednosti za ketamin, nije promjenjena uz BPC 157. Primjenom BPC 157 nisu promijenjene POA, postotak štakora s gubitkom refleksa, niti vrijeme oporavka refleksa uspravljanja. Primjenom BPC 157 poboljšana je somatosenzorna orijentacija nakon 50 minute (p<0,05 i p<0,001) i potencirano ubrzano kretanje štakora nakon 70-80 minute (p<0,001 i p<0,05). Fentanil je izazvao kataleptički odgovor u uskom rasponu doza, 2-3 minute nakon injekcije. Kumulativno trajanje katalepsije u štakora kojima je injiciran fentanil 300 μg/kg t. t. iznosilo je 22,23 ± 3,56 min. Katalepsija inducirana fentanilom bila je popraćena Straubovim fenomenom, opaženim kod mišićne rigidnosti. Primjena fentanila 200,0 μg/kg nije izazvala katalepsiju. Štakori kojima je primijenjen fentanil 400 µg/kg t.t. ispoljili su nevoljna uvrtanja trupa sa stražnjim šapama u abdukciji i letalnim ishodom nekolicine. Primjenom pentadekapeptida BPC 157 ng-μg značajno je skraćeno vrijeme oporavka katalepsije inducirano s fentanilom 300,0 μg/kg na 5,12 ± 1,29 i 4,15 ± 1,26 min (p<0,05). Vrijeme katalepsije skraćeno je + 76% za 10 ng/kg i 81% za 10 μg/kg BPC 157. Međutim, s BPC 157 se nije značajno utjecalo na poremećaj izazvan fentanilom 400 μg/kg. Droperidolom inducirana katalepsija trajala je duže od katalepsije izazvane fentanilom. Kataleptogeni učinak dehidrobenzperidola, ispoljen kod doze 6 mg / kg t. t., pojavo se tek u drugom satu i bio slabije izražen kod doze 8 mg / kg t. t. Primjenom pentadekapeptida BPC 157 10 μg ili 10 ng / kg t. t. oslabljena je katalepsiju u štakora tretiranih droperidolom sa 60,45 ± 2,25 na 0,00 ± 0,00 min (p < 0,05). Primjenom pentadekapeptida BPC 157 oslabljena je katalepsija kod niže doze (fentanil 0,10 mg/kg + droperidol 5,0 mg/kg) talamonala sa 20,00 ± 2,28 na 0,00 ± 0,00 min. Pentadekapeptid BPC 157 nema učinak na ponašanje normalnih životinja, ali je učinkovit u životinja kod kojih postoji poremećaj izazvan djelovanjem anestetika. Pokazano da BPC 157 smanjuje anestetski učinak tiopentala, odnosno hipnotski učinak diazepama, te skraćuje trajanje katalepsije izazvane fentanilom, droperidolom i talamonalom. Također, pokazano da BPC 157 ne mijenja anestetski učinak propofola i ketamina. BPC 157 ima učinke kod anestetika, u čijem je djelovanju posebno važna uloga NO/cGMP sustava, te posreduje biološke učinke i drugih neurotransmitera, čije djelovanje je povezano s njegovim primarnim učinkom. Sveukupno, rezultati upućuju na zaključak da BPC 157 ima antagonistički učinak na opće anestetsko djelovanje, te da postoji mogućnost da bi mogao poslužiti kao sredstvo za skraćenje vremena oporavka nakon prestanka djelovanja nekih anestetika.

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