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A simple method for short-term controlled anesthesia in newborn mice.

In this study, we describe a simple and inexpensive method for inducing short-term anesthesia and rapid recovery in newborn mice. Litters of Swiss mice pups were randomly allocated to testing on postnatal days 2, 5, and 8. Anesthesia was induced by placing the pup in a syringe and adding a volume of isoflurane-saturated gas that produced an estimated level of 32% isoflurane. Exposure to isoflurane lasted 30 s. All the pups survived the anesthesia. At all study ages, this method abolished the nociceptive response to tail clamp without inducing mortality, thus showing effective anesthesia. Recovery from anesthesia was assessed immediately after isoflurane exposure, based on two nonnoxious behavioral tests: the defensive response to a drop of water (10 tests, 1 min apart) and 10 min later the righting reflex, i.e., the time to recovery of the prone position (five tests, 10 min apart). The water drop test scores increased during the recovery phase toward the control values in all age groups. Treatment and time had no significant effect on righting reflex scores. The initial volume in the syringe, the volume of added isoflurane-saturated gas, and the duration of exposure may be adjusted according to postnatal age and specific strains or species (e.g., rats). This method is well suited to behavioral or physiological phenotype studies in developing mice, in which noxious procedures must precede functional testing, making rapid recovery from anesthesia a key requirement.

Age Factors↗

Kinetics of drug action in disease states. XXXI. Effect of experimental hyperthyroidism on the hypnotic activity of a benzodiazepine (oxazepam) in rats.

This investigation was designed to determine the effect of experimental hyperthyroidism on the hypnotic activity of a benzodiazepine and on the binding characteristics of the benzodiazepine receptor complex. Rats were made hyperthyroid by subcutaneous implantation of slow release pellets containing L-thyroxine. This treatment produced the characteristic symptoms of hyperthyroidism: increased serum thyroxine concentrations, increased heart weight and body temperature, and decreased serum protein concentrations. The hyperthyroid rats and a parallel group of normal animals (with drug-free pellets implanted subcutaneously) were slowly infused intravenously with oxazepam until they lost their righting reflex. The hyperthyroid rats required a significantly larger dose of the benzodiazepine and their total serum and cerebrospinal fluid concentrations of oxazepam (but not the serum concentration of free drug and the brain concentration) at the onset of loss of the righting reflex were modestly but statistically significantly lower than those of the normal rats. The receptor density and affinity for diazepam in hyperthyroid rats were not significantly different from those of the normal animals. Hyperthyroidism apparently affects the pharmacokinetics of oxazepam but has, at best, only a small effect on the pharmacodynamics (hypnotic activity) of the drug.

Animals↗

In rats breathing from a nonrebreathing system, substitution of desflurane for isoflurane toward the end of anesthesia incompletely restores the time of recovery toward that of desflurane.

UNLABELLED: The lower solubility of desflurane allows a more rapid emergence from anesthesia than after anesthesia with the more soluble but less expensive anesthetic, isoflurane. Some practitioners use isoflurane for maintenance of anesthesia, crossing over to desflurane later in maintenance in an attempt to combine the cost-effectiveness of isoflurane with the rapid emergence from desflurane. We hypothesized that this maneuver would not accomplish its goals. Twenty-four male Sprague-Dawley rats received 1.2 minimum alveolar anesthetic concentration (MAC) of desflurane for the final 15, 30, or 60 min of a 2-h, 1.2-MAC isoflurane anesthetic in a nonrebreathing anesthesia system. We measured the time from cessation of anesthetic administration to the time each rat righted himself twice. Immediately after righting for the second time, we tested each rat's ability to remain atop a rotating rod (Rota-Rod) for 60 s continuously. Early (righting reflex) and late (Rota-Rod) recovery occurred more rapidly (P < 0.001) after 120 min of anesthesia with desflurane alone than after 120 min of anesthesia with isoflurane alone. A cross-over period of 30 min or longer produced a righting reflex time that did not differ from that found with desflurane alone, but a 15-min cross-over did not. Progressively longer cross-over periods led to proportionally better Rota-Rod performance, but no cross-over duration produced the rapidity of recovery seen with desflurane alone. We concluded that in a nonrebreathing system, switching to desflurane during the last 30 min of anesthesia substantially improved early recovery but produced a much smaller improvement in later recovery. IMPLICATIONS: The newer inhaled anesthetics offer the advantage of lower solubility, and thus more rapid emergence from anesthesia, than do the older inhaled anesthetics. However, they can be more expensive to use. This study demonstrates that substituting the newer anesthetic, desflurane, toward the end of anesthesia for an older anesthetic of greater solubility, isoflurane, does not produce recovery comparable to that of desflurane alone. Furthermore, this technique can be more costly than using desflurane throughout anesthesia.

Anesthetics, Inhalation↗

Behavioral and neuroanatomical sequelae of prenatal naloxone administration in the rat.

Pregnant Long-Evans hooded rats were dosed subcutaneously with 1 or 5 mg/kg/day naloxone hydrochloride, or an equal volume of vehicle, from gestational Day 4 (GD4) through GD19. Offspring were assessed for development of righting reflex, negative geotaxis, and open field activity, and for acquisition of a Warden maze; offspring sacrificed at postnatal Day (PND) 21 were assessed for several parameters of cerebellar, hippocampal, and motor cortical morphology. Five mg/kg/day naloxone accelerated development of negative geotaxis and righting reflex, while 1 mg/kg/day naloxone tended to slow development. Low dose females had significantly more errors than controls on the first day of maze learning. The high dose group had a significantly higher concentration of granule cells in the curvature of the dentate gyrus than controls; other neuroanatomical measures were unaffected by dosing. These findings confirm and extend previous work indicating that prenatal exposure to naloxone may alter neurobehavioral development in the rat.

Animals↗

Effects of chronic alcohol intake on anesthetic responses to diazepam and thiopental in rats.

The effect of chronic alcohol intake on anesthetic responses to alcohol, thiopental, or diazepam was examined in adult male Sprague-Dawley rats. Alcohol-fed animals were maintained solely on a complete balanced liquid diet containing 6.54% ethanol (w/w) for 21 days; pair-fed control animals received equal amounts of the same diet with alcohol isocalorically replaced by sucrose or dextrin. Nine hours after diets were withdrawn on the twenty-second day, the following drug/dose combinations were administered intraperitoneally to separate groups of alcohol-fed and control rats (10-15 animals in each group): ethanol 2.4, 3.2, and 4.0 g/kg; thiopental 20, 40, and 80 mg/kg; and diazepam 10, 20, and 40 mg/kg. Three different responses were assessed in every animal: 1) loss of righting reflex (induction of anesthesia); 2) response to a painful stimulus (analgesia); and 3) sleeping time (duration of anesthesia). Alcohol-fed rats compared with controls were significantly less tolerant of pain at an acute alcohol dose of 2.4 g/kg, and loss of righting reflex and sleeping time were reduced at 4.0 g/kg. All three anesthetic responses were also attenuated in alcohol-fed rats at a diazepam dose of 20 mg/kg. In contrast, none of the three responses was reduced in alcohol-fed rats at any of the three thiopental doses. Thus, chronic alcohol intake sufficient to produce tolerance to anesthetic doses of alcohol in rats also produced cross-tolerance to diazepam but not to thiopental in equianesthetic doses. These results suggest that blanket recommendations for adjusting intravenous anesthetic dosages in alcoholic humans may be inadequate as guides to anesthetic management.

Alcoholism↗

Effect of flumazenil in a model of acute alcohol intoxication in rats.

The ability of high doses of flumazenil to antagonize ethanol-induced sedation was assessed via ethanol-induced loss of the righting reflex (LRR) in rats. Ethanol (3.5-5 g kg-1 i.p) produces a dose-dependent increase in the proportion of animals presenting a LRR response. Flumazenil (0.1 to 1 mg kg-1 i.p) was ineffective in preventing the suppression of the righting reflex and even aggravated it for ethanol dosages of 3.5 and 4.0 g kg-1. These data support the view that flumazenil has no effective 'anti-ethanol' properties. Conversely, the prolongation of mean sleep time provoked by the antidote could be related to the suppression of a putative endogenous inverse-agonist activity at the benzodiazepine receptor, which could be implicated in the mechanism of the acute tolerance to ethanol.

Alcoholic Intoxication↗

Neurotoxic effects of phenytoin on postnatal mouse brain development following neonatal administration.

Phenytoin (PHT) is a commonly used anticonvulsant drug. It has been reported that children exposed prenatally to PHT have brain malformations and psychomotor dysfunction. The neonatal development of the central nervous system (CNS) in mice corresponds to the last trimester in humans. To examine the neurotoxic effects of PHT on postnatal brain development, we administered PHT at doses of 10, 17.5, 25, or 35 mg/kg to newborn mice once a day during postnatal days (PD) 2-4. These dose levels result in plasma levels corresponding to the therapeutic ranges in humans. We measured the weight of total brain, cerebrum, cerebellum, and brain stem on PD 5 through 21, and examined early motor functions including head elevation, elevation of pelvis, pivoting, crawling, and righting reflex . Total brain weight, cerebral weight, and cerebellar weight in the group treated with 25 or 35 mg/kg were significantly reduced compared to controls from PD 5 to 21. Mice treated with PHT at 25 or 35 mg/kg showed decreased locomotor abilities and righting reflex on PD 5. In all phenytoin treatment groups, phenytoin levels in the brain were higher than those in the plasma on the third day of PHT treatment. We thus observed neurotoxic effects of PHT on postnatal brain development in mice. Our present data may provide useful implications for the management of PHT-induced developmental neurotoxicity and evaluation of psychomotor development in children exposed to PHT during the late fetal period.

Aging↗

Decreased ethanol preference and consumption in dopamine transporter female knock-out mice.

BACKGROUND: It is commonly believed that the mesolimbic dopamine (DA) system participates in the etiology of alcoholism. One of the most important regulators of DA synaptic transmission is the DA transporter (DAT). We examined the effects of the genetic reduction or deletion of DAT on voluntary ethanol consumption and ethanol-induced sedation. METHODS: Ethanol preference and consumption were assessed in the two-bottle choice paradigm, and the depressant effects of ethanol were evaluated by measuring sleep time after acute injection of ethanol. RESULTS: The latent period to lose the righting reflex was shorter in both knock-out mice (DAT-KO) and heterozygote mice (HET) than in wild-type (WT) mice. No significant difference was found among the three genotypes in the ethanol blood concentration at the onset of regaining the righting reflex. Females of all genotypes consumed more fluid than males of the same genotype. HET and DAT-KO females had increased total fluid consumption compared with WT females. DAT-KO males had increased fluid consumption compared with WT and HET males. Ethanol preference and consumption were not different among male mice of different genotypes. WT and HET females demonstrated significantly higher ethanol consumption than males. HET female mice did not differ from WT mice in ethanol preference. There was no difference between HET and WT mice in the preference for saccharin or quinine solutions. DAT-KO females avoided ethanol, and their consumption and preference were lower than in WT and HET females, despite markedly increased total intake. DAT-KO mice also demonstrated altered taste preference for saccharin and quinine. CONCLUSIONS: Partial deletion of DAT results in increased fluid consumption in female mice but does not change ethanol preference in either sex. Complete deletion of DAT reduces ethanol preference in female mice; this may be due to a combination of the pharmacological actions of DAT deletion and alterations in fluid consumption and taste discrimination.

Alcohol Drinking↗

Regional distribution of barbital in the brain of mice during the development of tolerance and physical dependence.

ICR male mice each receiving a 16 mg barbital pellet implanted subcutaneously for three days developed about 40 percent tolerance to barbital and more than 50 percent tolerance to pentobarbital as measured by sleeping time. The development of physical dependence in these mice was demonstrated by an increased sensitivity to convulsions with pentylenetetrazol. The concentration of barbital rose to high levels in the subcortex and the cerebellum during the time between the implantation of the pellet and the loss of the righting reflex and distributed uniformly thereafter. After a challenge dose of the drug, higher barbital concentrations were found in these two areas as well as in the pons-medulla at 30 minutes and in all areas at the time of the loss of righting reflex, in withdrawn mice. The latter finding would indicate an increase in threshold for barbital anesthesia. The mechanisms of the higher uptake in the brain of withdrawn mice are discussed.

Animals↗

Changes in gravity influence rat postnatal motor system development: from simulation to space flight.

Our research examines the role of the environment in postnatal nervous system development. Recently we have been studying the effects of changes in gravity on the motor system of rats from postnatal day (P) 2 to 31 using kinematic analysis of swimming, walking, and righting reflexes. Using the tail suspension model of weightlessness we identified sensitive and critical periods of motor system development corresponding to the time during which a motor skill is first achieved. Motor performance in suspended animals was marked by slow swimming, walking, and air-righting, all of which were characterized by hindlimb extension. (Walton et al, Neurosci. 52,763,1992). The critical periods identified in these studies contributed to determining the age of animals for a small payload, NIH.R3. This 9-day mission (STS-72) included 2 litters at P5, P7, or P15 at launch. The P7-16 and P15-24 groups were studied post-flight. On the landing day (R+0) surface righting, swimming and walking were slower in flight compared to control animals. Differences were more marked in the younger animals and the hindlimbs were more affected than the forelimbs with marked, prolonged extension of, at least, the ankle joint angle. Readaptation to 1G was slower in the P7-16 group with righting reflexes adapting first, walking last. We have shown that gravity is an important factor in postnatal nervous system development and that its affect depends on the age of the animal, duration of the perturbation, and the motor function studied.

Animals↗

Perinatal caffeine treatment: behavioral and biochemical effects in rats before weaning.

Administration of drinking water containing 0, 0.02%, 0.04% and 0.08% of caffeine to female rats throughout gestation and lactation affects several behavioral parameters in the offspring. Righting reflexes, swimming ability development, motor coordination and muscle tone were affected. The activity of these animals, as measured with an open-field test at weaning (i.e., at the end of the treatment), was reduced. The effects observed were dose-dependent. However, for righting reflexes the dose-dependency was direct (the highest dose producing maximal effects), whereas in all the other tests, the dose-dependency was inverse, the lowest dose producing maximal effects and the highest dose producing no effects. This might reflect the presence of subclasses of receptors having different affinities for adenosine, mediating opposite effects and antagonized by caffeine. On the other hand, perinatal caffeine effects are certainly not mediated by blockade of phosphodiesterases, since cAMP levels at the end of the treatment were dose-dependently reduced. This study shows therefore that administration of caffeine to rat dams is able to influence the neurobehavioral development of the offspring. Moreover, all the doses utilized and corresponding to 27, 58 and 108 mg/kg, were able to produce all or some of the mentioned effects, indicating that further testing with doses lower than 27 mg/kg is required to find a dose which does not affect the offspring.

Animals↗

Precipitated withdrawal in squirrel monkeys after repeated daily oral administration of alprazolam, diazepam, flunitrazepam or oxazepam.

The lowest dose of alprazolam, diazepam, flunitrazepam and oxazepam consistently to induce loss of righting reflex in squirrel monkeys or vehicle was orally administered to monkeys on 18 consecutive days: 2 mg/kg alprazolam (n = 4), 30 mg/kg diazepam (n = 4), 1 mg/kg flunitrazepam (n = 4), 280 mg/kg oxazepam (n = 5), or vehicle (n = 4). Tolerance developed rapidly for loss of righting reflex, more slowly for sleep and only minimally for muscle relaxation observed during the period immediately following daily oral administration. Injection of the specific benzodiazepine receptor antagonist flumazenil (10 mg/kg i.v.) 5 h after the ninth daily oral treatment produced signs of precipitated withdrawal (tremor, vomiting and/or convulsions) in one alprazolam-, four diazepam-, one flunitrazepam- and four oxazepam-treated monkeys, but not in the vehicle-treated monkeys. Physiological saline injected intravenously several days later under these same experimental conditions failed to provoke a precipitated withdrawal reaction. When flumazenil-induced precipitated withdrawal was again evaluated after the 18th daily oral treatment, withdrawal signs were observed in all alprazolam- and all diazepam-treated monkeys, as well as in three flunitrazepam- and three oxazepam-treated monkeys, but not in the vehicle-treated monkeys (convulsions were observed in one alprazolam-, two diazepam-, one flunitrazepam- and two oxazepam-treated monkeys). No signs of spontaneous withdrawal were observed in any of the monkeys during a subsequent 3-week drug-free period. Thus, repeated administration of approximately equieffective doses of these four benzodiazepines resulted in a similar development of tolerance and physical dependence (indicated by the occurrence of a precipitated withdrawal reaction).

Administration, Oral↗

Developmental age influences the effect of epidural dexmedetomidine on inflammatory hyperalgesia in rat pups.

BACKGROUND: Epidural alpha2-adrenergic agonists produce analgesic effects in children and adults, but efficacy and safety have not been established in neonates and infants. The aim of this study was to determine the effect of epidural dexmedetomidine on sensory processing, reversal of inflammatory hyperalgesia, and sedation during early development in rats. METHODS: In rat pups aged 3, 10, and 21 postnatal days, mechanical withdrawal thresholds of the hind limbs were measured at baseline and after unilateral inflammation due to carrageenan. The effect of epidural dexmedetomidine on withdrawal thresholds was measured for 90 min after injection, and dose-response curves were constructed for each age group. The duration of the righting reflex was measured to assess sedation. The effects of epidural and systemic administration of dexmedetomidine were compared. RESULTS: At all ages, carrageenan-induced hyperalgesia was reversed by doses of epidural dexmedetomidine that did not affect the threshold of the contralateral paw or prolong the righting reflex. Higher doses of epidural dexmedetomidine affected baseline nociception in the contralateral paw and produced sedation but had no effect when given systemically. Reversal of hyperalgesia and sedation were produced by lower doses of epidural dexmedetomidine in the youngest pups. CONCLUSIONS: Spinally mediated selective reversal of inflammatory hyperalgesia by epidural dexmedetomidine can be achieved at all ages; relatively lower doses are effective in early life, but the therapeutic window is narrow. These data have implications for the use and dosing of epidural alpha2 agonists in neonates and infants.

Age Factors↗

Catalepsy induced by combinations of ketamine and morphine: potentiation, antagonism, tolerance and cross-tolerance in the rat.

Previous studies demonstrated that both ketamine and morphine induced analgesia and catalepsy in the rat. Pre-treatment with ketamine produced cross-tolerance to morphine, whereas pretreatment with morphine did not induce cross-tolerance to ketamine but rather augmented the cataleptic response; this augmentation was attributed to residual morphine in the brain. The present studies explored the duration of the loss of righting reflex induced by sub-effective doses of ketamine and morphine, administered simultaneously. There was mutual potentiation between sub-effective doses of ketamine and morphine, but sub-effective doses of ketamine partly antagonized fully-effective doses of morphine. Latency to the loss of righting reflex, rigidity and behavior on recovery, reflected the relative predominance of ketamine or morphine in each combination. Naloxone inhibited the induced cataleptic effects. The degree and time course of development of tolerance to daily administration of sub-effective dose combinations of ketamine and morphine were similar. Rats, tolerant to ketamine-dominant combinations, were cross-tolerant to both drugs, while those tolerant to morphine-dominant combinations were cross-tolerant to morphine but showed either no cross-tolerance or an augmented response to ketamine. While the mutual potentiation, antagonism and tolerance suggest common mechanisms for the induced catalepsy, differences in latency, rigidity and behavior, asymmetry of cross-tolerance and a widely-different ID50 for naloxone would argue against an action at a single opioid site.

Animals↗

Actions of long chain alcohols on GABAA and glutamate receptors: relation to in vivo effects.

1. The effects of n-alcohols on GABAA and glutamate receptor systems were examined, and in vitro effectiveness was compared with in vivo effects in mice and tadpoles. We expressed GABAA, NMDA, AMPA, or kainate receptors in Xenopus oocytes and examined the actions of n-alcohols on receptor function using two-electrode voltage clamp recording. 2. The function of GABAA receptors composed of alpha 1 beta 1 or alpha 1 beta 1 gamma 2L subunits was potentiated by all of the n-alcohols studied (butanol-dodecanol). 3. In contrast to GABAA receptors, glutamate receptors expressed from mouse cortical mRNA or from cRNAs encoding AMPA (GluR3)- or kainate (GluR6)-selective subunits were much less sensitive to longer chain alcohols. In general, octanol and decanol were either without effect or high concentrations were required to produce inhibition. 4. In contrast to the lack of behavioural effects by long chain alcohols reported previously, decanol produced loss of righting reflex in short- and long-sleep mice, indicating that the in vivo effects of decanol may be due in part to actions at GABAA receptors. Furthermore, butanol, hexanol, octanol, and decanol produce similar potentiation of GABAA receptor function at concentrations required to cause loss of righting reflex in tadpoles, an in vivo model where alcohol distribution is not a compromising factor. 5. Thus, the in vivo effects of long chain alcohols are not likely to be due to their actions on NMDA, AMPA, or kainate receptors, but may be due instead to potentiation of GABAA receptor function.

Alcohols↗

Separate site(s) of action of optical isomers of 1-methyl-5-phenyl-5-propylbarbituric acid with opposite pharmacological activities at the GABA receptor complex.

The behavioral profile of the optical isomers of 1-methyl-5-phenyl-5-propylbarbituric acid (MPPB) and their interaction with the convulsant binding site at the GABA receptor complex were investigated. R(-)-MPPB produced dose-related loss of righting reflex, whereas S(+)MPPB produced convulsions in a dose-dependent manner. Subconvulsive doses of S(+)MPPB were proconvulsant with a subeffective dose of picrotoxin. S(+)MPPB-induced seizures were blocked by R(-)MPPB and pentobarbital. In contrast, S(+)MPPB did not block the loss of righting reflex produced by R(-)MPPB or pentobarbital. S(+)MPPB inhibited the binding of [35S]t-butylbicyclophosphorothionate (TBPS), a ligand that binds to the picrotoxin site on the oligomeric GABA receptor complex, to rat brain membranes competitively, whereas R(-)MPPB inhibited it noncompetitively. Thus, the optical isomer of MPPB, which have opposite pharmacological effects, interact differently with the convulsant (TBPS) site at the GABA receptor complex. These results suggest that the convulsant S(+)MPPB and the depressant R(-)MPPB may produce their behavioral effects by acting via convulsant and anticonvulsant sites, respectively, at the GABA receptor complex.

Animals↗

The lack of effect of opioid agonists and antagonists on some acute effects of ethanol.

We studied the ability of opioid antagonists: naloxone, naltrexone and diprenorphine and an opioid agonist morphine to influence the effects of ethanol on hypothermia, sleeping time and impairment of aerial righting reflex. Naltrexone (2-16 mg/kg) and naloxone (2-16 mg/kg) were not able to attenuate effects of ethanol, while diprenorphine decreased ethanol sleeping time (4 microgram/kg) and antagonized the ethanol hypothermia (8 microgram/kg). Naltrexone in a dose of 8 mg/kg sc antagonized the ethanol impairment of aerial righting reflex. The present behavioral studies did not provide any evidence for the participation of the opioid system in the mediation of acute ethanol effects in rats.

Animals↗

Lumbomuscular proprioceptive reflexes in body equilibrium.

To evaluate the role of reflexes related to the lumbar proprioceptors in maintenance of body equilibrium, changes in equilibrium function of the eyes and body were observed after unilateral procainization of the lumbar erector muscles. Observations were made on normal subjects and vertigo cases with lumbar pain after whiplash injury using various equilibrium tests. The results obtained were as follows: (1) On unilateral procainization of the lumbar erector muscles of normal subjects, eye nystagmus and disturbances of the righting reflex developed. Simultaneously, changes in drift reactions of the lower limbs were detected by the stepping test. Namely, in many of the subjects examined the direction of stepping deviation became quite different from that before procainization, and stepping after procainization tended to show slight or moderate ataxic features, associated with a sensation of unsteadiness. (2) When procaine was injected unilaterally into tender spots in the lumbar erector muscles of traumatic vertigo cases, spontaneous eye nystagmus and disturbances of the righting reflex decreased. Simultaneously, significant changes in the drift reactions of the lower limbs were observed in many of the cases examined. Namely, the direction of deviation became the opposite of that before procainization and ataxia in walking almost disappeared with reduction in vertigo. The following conclusions were drawn from these findings: (1) The effects of procaine on equilibrium of normal subjects are in sharp contrast to its effects on equilibrium of traumatic vertigo cases. Findings in the former might be due to increased imbalance between the activities of the right and left lumbar proprioceptors, while those in the latter might be due to decreased imbalance between the two. (2) These findings support the view that from the standpoint of body equilibrium, there are two phases of the proprioceptive reflex, and that Fukuda's concept of "two phases of the labyrinthine reflex, i.e., a stage of disturbance and a stage of coordination", can be applied to interpretation of the proprioceptive reflex of lumbar origin.

Adult↗