Joseph R. Tobin

Dexmedetomidine-Induced Sedation in Volunteers Decreases Regional and Global Cerebral Blood Flow

Dexmedetomidine is a selective &agr;2-agonist approved for sedation of critically ill patients. There is little information on the effects of dexmedetomidine on cerebral blood flow (CBF) or intracranial hemodynamics, despite considerable other pharmacodynamic data. We hypothesized that therapeutic doses of dexmedetomidine would decrease CBF. Therefore, nine supine volunteers, aged 24–48 yr, were infused with a 1 &mgr;g/kg IV loading dose of dexmedetomidine, followed by an infusion of 0.2 &mgr;g · kg−1 · h−1 (LOW DEX) and 0.6 &mgr;g · kg−1 · h−1 (HIGH DEX). Hemodynamic and CBF (via positron emission tomography) measurements were determined at each experimental time point. Dexmedetomidine decreased both cardiac output and heart rate during and 30 min after drug administration. Blood pressure decreased from 12% to 16% during and after the dexmedetomidine administration. Global CBF was decreased significantly from baseline (91 mL · 100 g−1 · min−1 [95% confidence interval, 72–114] to 64 mL · 100 g−1 · min−1 [51–81] LOW DEX and 61 mL · 100 g−1 · min−1 [48–76] HIGH DEX). This decrease in CBF remained constant for at least 30 min after the dexmedetomidine infusion was discontinued, despite the plasma dexmedetomidine concentration decreasing 40% during this same time period (628 pg/mL [524–732] to 380 pg/mL [253–507]).

Glutamate-Induced Cerebral Vasodilation Is Mediated by Nitric Oxide Through N-Methyl-d-Aspartate Receptors

BACKGROUND AND PURPOSE It was found that glutamate, a major neurotransmitter, is vasoactive in the cerebral circulation. However, the mechanism is unclear. This study was designed to investigate the role of nitric oxide (NO) and N-methyl-D-aspartate (NMDA) receptors in cerebral arteriolar dilation to glutamate. METHODS Newborn, chloralose-anesthetized pigs were equipped with a closed cranial window. The diameter of pial arterioles was measured by means of intravital microscopy, and NO synthase (NOS) activity in brain cortex was determined by the conversion assay of [14C]arginine to [14C]citrulline. RESULTS Topical application of glutamate at 10(-7), 10(-6), and 10(-5) mol/L (n = 5) increased the mean diameter by 12 +/- 3%, 13 +/- 2%, and 18 +/- 3% (+/- SEM), respectively (baseline, 91 +/- 10 microns; P < .05). Similarly, NMDA application at the above doses (n = 5) dilated arterioles by 10 +/- 2%, 16 +/- 3%, and 18 +/- 6%, respectively (baseline, 97 +/- 4 microns; P < .05). Topical application of 10(-4) mol/L NG-nitro-L-arginine (L-NNA), which inhibited NOS activity by 93%, blocked the arteriolar dilation to glutamate or NMDA. Furthermore, administration of MK-801, a potent inhibitor of NMDA receptors, blocked glutamate-induced vasodilation completely in both topical application (10(-5) mol/L; n = 6) and intravenous administration (5 to 10 mg/kg; n = 5). In addition, neither L-NNA nor MK-801 attenuated the vasodilation to hypercapnia (PCO2 = 40 to 68 mm Hg). CONCLUSIONS Glutamate-induced cerebral arteriolar dilation is mediated by NO through NMDA receptors, and NO does not play a major role in the cerebral arteriolar dilation to hypercapnia (PCO2 = 40 to 68 mm Hg) in newborn pigs.

Screening of the entire ryanodine receptor type 1 coding region for sequence variants associated with Malignant hyperthermia susceptibility in the North American Population

Background:Malignant hyperthermia (MH) is a life-threatening and frequently fatal disorder triggered by commonly used anesthetics. MH susceptibility is a genetically determined predisposition to the development of MH. Mutations in the ryanodine receptor type 1 (RYR1) gene are the major cause of MH susceptibility. The authors sought to develop a reliable genetic screening strategy based on efficient and relatively inexpensive mutation-detection procedures. Methods:A cohort (n = 30) of North American MH patients and MH-susceptible individuals was studied. RNA and DNA extracted from muscle tissue or blood lymphocytes were used for analysis. The entire RYR1 coding region was amplified in 57 overlapping fragments and subjected to denaturing high-performance liquid chromatography analysis followed by direct nucleotide sequencing to characterize RYR1 alterations. Results:Nine previously reported and nine unknown RYR1 mutations were identified in 21 of 30 studied patients (70%). Some of the new mutations were located outside of known mutational “hot spots,” suggesting that RYR1 contains previously unknown mutation-prone areas requiring analysis. The North American MH/MH-susceptible population is characterized by a high RYR1 allelic heterogeneity. Conclusions:Denaturing high-performance liquid chromatography analysis of RNA samples extracted from the biopsied skeletal muscle followed by DNA sequencing is a highly efficient methodology for RYR1 mutation detection. This approach allows increasing the rate of mutation detection to 70% and identifying mutations in the entire RYR1 coding region.

Bacterial Colonization and Infection Rate of Continuous Epidural Catheters in Children

Continuous epidural infusions are widely used for postoperative analgesia in children.We prospectively studied the incidence of bacterial colonization of caudal and lumbar epidural catheters, as well as the incidence of serious systemic and local infection, in 210 children after short-term epidural analgesia. Using aseptic technique, epidural catheters were inserted into either the lumbar or the caudal epidural space based on the preferences of the anesthesia team and/or clinical indication. The integrity of the catheter and overlying transparent dressing site was evaluated by a member of the pediatric pain service at least once a day. The catheters were aseptically removed if the patient had a fever greater than 39[degree sign]C, if the dressing was compromised, or when epidural analgesia was no longer required. The subcutaneous portion of the catheter was semiquantitatively cultured. Cellulitis (erythema, swelling, purulent discharge, pustule formation, or tenderness) was diagnosed by examination of the epidural insertion site. The mean (+/- SD) age of patients in the caudal catheter group (n = 170) was 3 +/- 3 yr; their mean weight was 13 +/- 11 kg. The mean (+/- SD) age of patients in the epidural catheter group (n = 40) was 11 +/- 4 yr; their mean weight was 36 +/- 23 kg. All catheters remained in place for 3 +/- 1 days (range 1-5 days). There was no serious systemic infection (meningitis, epidural abscess, or systemic sepsis). Of all epidural catheters, 35% (73 of 210) were colonized. Gram-positive colonization was similar in caudal (25%; 43 of 170) and lumbar (23%; 9 of 40) catheters. Gram-negative organisms were cultured from 16% of the caudal catheters (27 of 170) and 3% of the lumbar catheters (1 of 40). In patients treated with caudal epidural catheters, children aged >3 yr were less likely to have colonized epidural catheters than younger children. Age did not affect the probability of developing cellulitis at the insertion site. Although patients aged <3 yr with caudal catheters had a slightly greater risk of cellulitis than children aged >3 yr (14% vs 9%), this association was very weak (P = 0.33). We observed that, despite bacterial colonization of caudal and lumbar epidural catheters, serious systemic and local infection after short-term epidural analgesia did not occur in our study. Implications: Continuous epidural infusions are widely used for postoperative analgesia in children. We found no serious systemic infections after short-term (3 days) continuous epidural analgesia in children. (Anesth Analg 1998;86:712-6)

Neuropathic pain in rats is associated with altered nitric oxide synthase activity in neural tissue.

Peripheral nerve injury may lead to a chronic neuropathic pain state that results from an increase in excitability of central neurons. This central sensitization is mediated via an N-methyl-D-aspartic acid (NMDA) receptor and may involve the production of nitric oxide (NO). As NO is suggested to play a role in nociceptive transmission following nerve injury, we examined for altered NO synthase activity at multiple levels of peripheral and spinal neural tissue in a rat model of neuropathic pain. Peripheral neuropathy was induced in rats (N = 12) by ligation of the left L5 and L6 nerve roots. Six other rats had sham surgery. An ipsilateral decrease in paw withdrawal threshold to mechanical stimuli confirmed the presence of a neuropathic pain state. Samples of the lumbar and thoracic spinal cords, L4, L5, and L6 dorsal root ganglia (DRGs), and the sciatic nerves were obtained from the lesioned and contralateral sides at 2 and 4 weeks after neuropathic surgery (N = 6 per group). In the lumbar spinal cord, a bilateral decrease in nitric oxide synthase (NOS) activity was observed 2 and 4 weeks after neuropathic surgery. NOS activity was increased in the ipsilateral L5 and 6 DRGs 2 weeks following neuropathic surgery. An increase in NOS activity in the DRG may be an early mechanism for inducing more central changes. The bilaterally decreased NOS activity in the lumbar spinal cord may be secondary to a negative feedback mechanism resulting from increased NO production in the spinal dorsal root ganglia. Multiple alterations in expression of NOS activity that occur in both peripheral and central processing may play a role in the pain behavior resulting from peripheral nerve injury. (Preliminary results of these studies have been presented in abstract form at the annual meetings of the Society for Neuroscience, 1994, and the American Society of Anesthesiologists, 1994).

Sex differences in cholinergic analgesia I: a supplemental nicotinic mechanism in normal females.

BACKGROUND Cholinergic agents produce analgesia after systemic and intrathecal administration. A retrospective review showed that intrathecal neostigmine was more potent in women than in men, suggesting a sex difference in this response. The purpose of this study was to determine whether such a sex difference exists in normal rats and to examine the pharmacologic mechanisms that underlie this difference. METHODS Male and female rats with indwelling intrathecal catheters received injections of neostigmine, bethanechol (muscarinic agonist), or RJR-2403 (neuronal nicotinic agonist) alone or with atropine (muscarinic antagonist), mecamylamine (nicotinic antagonist), or phentolamine alpha-adrenergic antagonist) with antinociception determined to a noxious heat stimulus to the hind paw. Time versus subcutaneous paw temperature relationships were defined for males and females. RESULTS Neostigmine produced dose-dependent antinociception with five times greater potency in female than in male rats. Neostigmine-induced antinociception was reversed in male rats by atropine and unaffected by mecamylamine, whereas it was partially reduced by each antagonist alone in females and completely reversed after injection of both. RJR-2403 was more potent in females than in males, whereas there was no sex difference to bethanechol. Phentolamine partially reversed antinociception from RJR-2403 in females. Paw temperature increased more rapidly in females than in males for the same lamp intensity. CONCLUSIONS These data demonstrate a large sex difference in antinociception to intrathecal neostigmine that is primarily the result of a nicotinic component in females. Phentolamine reversal suggests that part of this nicotinic component may rely on spinal norepinephrine release. A better understanding of this sex difference could lead to development of novel pain therapy for women.

Selective anesthetic inhibition of brain nitric oxide synthase.

BackgroundIt has been postulated that nitric oxide (NO) is a neurotransmitter involved in consciousness, analgesia, and anesthesia. Halothane has been shown to attenuate NO-mediated cyclic guanosine monophosphate accumulation in neurons, and a variety of anesthetic agents attenuate endothelium-mediated vasodilation, suggesting an Interaction of anesthetic agents and the NO-cyclic guanosine monophosphate pathway. However, the exact site of anesthetic inhibitory action in this multistep pathway is unclear. The current study examines effects of volatile and Intravenous anesthetic agents on the enzyme nitric oxide synthase (NOS) in brain. MethodsNOS activity was determined by in vitro conversion of [14C]arginine to [14C]citrulline. Wistar rats were decapitated and cerebellum quickly harvested and homogenized. Brain extracts were then examined for NOS activity in the absence and presence of the volatile anesthetics halothane and isoflurane, and the intravenous agents fentanyl, midazolam, ketamine, and pentobarbltal. Dose-response curves of NOS activity versus anesthetic concentration were constructed. Effects of anesthetics on NOS activity were evaluated by analysis of variance. ResultsControl activities were 57.5 ± 4.5 pmol · mg protein-1 min-1 in the volatile anesthetic experiments and 51.5 ± 6.5 pmol · mg protein-1 · min-1 in the intravenous anesthetic experiments. NOS activity was not affected by ketamine (≤I × × 10-4 M), pentobarbital (≤5 × 10-5 M), fentanyl (≤1 × 10-5 M), and midazolam (≤1 × 10-5 m). Halothane decreased NOS activity to 36.7 ± 2.5 (64% of control, P < 0.01 from control), 23.8 ± 4.3 (41%, P < 0.01 from control and <0.05 from 0.5% halothane), 25.2 ± 3.8 (44%, P < 0.01 from control and <0.05 from 0.5% halothane), and 19.7 ± 2.8 (34%, P < 0.01 from control and <0.05 from 0.5% halothane) pmol · mg protein-1 · min-1 at 0.5,1.0,2.0, and 3.0% vapor. Isoflurane decreased NOS activity to 48.9 ± 6.1 (85% of control), 46.0 ± 3.2 (80%, P < 0.05 from control), 40.3 ± 5.1 (70%, P < 0.05 from control), and 34.2 ± 4.0 (60%, P < 0.05 from control and 0.5% and 1.0% isoflurane) pmol · mg protein-1 · min-1 at 0.5,1.0,1.5, 2.0% vapor, respectively. ConclusionsVolatile anesthetics inhibit brain NOS activity in an in vitro system, but the Intravenous agents examined have no effect at clinically relevant concentrations. This inhibition suggests a protein-anesthetic interaction between halothane, isoflurane, and NOS. In contrast, intravenous agents appear to have no direct effect on NOS activity. Whether intravenous agents alter signal transduction or regulatory pathways that activate NOS is unknown.

Effects of two years of estrogen loss or replacement on nucleus basalis cholinergic neurons and cholinergic fibers to the dorsolateral prefrontal and inferior parietal cortex of monkeys

The present study examined the long‐term (2 years) effects of estrogen loss or estrogen replacement therapy (ERT) on cholinergic neurons in the nucleus basalis of Meynert and on cholinergic fibers in the prefrontal and parietal cortex of adult female cynomolgus monkeys. Cholinergic fiber density in layer II of the prefrontal cortex was decreased in monkeys who were ovariectomized and treated with placebo for 2 years. In contrast, ovariectomized monkeys receiving ERT for 2 years had fiber densities that were comparable to those of intact controls. No differences in parietal cholinergic fiber density or nucleus basalis cholinergic neuron number or volume were found among intact, ovariectomized, or ERT monkeys. Our results suggest that ERT is effective in preventing region‐specific changes in cortical cholinergic fibers that result from the loss of circulating ovarian hormones. These modest but appreciable effects on cholinergic neurobiology following long‐term estrogen loss and ERT may contribute to changes in visuospatial attention function that is mediated by the prefrontal cortex. J. Comp. Neurol. 469:507–521, 2004.

Age-dependent Responses to Thermal Hyperalgesia and Mechanical Allodynia in a Rat Model of Acute Postoperative Pain

Background Developmental differences in short- and long-term responses to pain, especially surgical pain, have received minimal attention. The purpose of the present study was to examine postoperative responses in rats of developmental ages paralleling the infant to young adult human. Methods The withdrawal threshold to von Frey filament testing and withdrawal latency to hind-paw radiant heating were determined before and for various times after hind-paw incision in rats 2, 4, and 16 weeks of age. Control rats of these ages were observed serially without surgery. Results In control animals, younger rats were more sensitive to mechanical stimulation and less sensitive to thermal stimulation. Paw incision resulted in similar changes to both types of stimulation in all age groups, peaking 4 h after surgery. However, the return to normal sensitivity to mechanical stimulation, as measured by return of threshold to 80% of normal, occurred more quickly in 2-week-old than in 4- and 16-week-old animals. In contrast, there was no age difference for time to return to normal sensitivity to thermal stimulation after surgery. Conclusions The more rapid recovery of the younger animals from the mechanical allodynia but not thermal hypersensitivity after surgery suggests the presence of developmental differences in modulation of A-fiber sensitization after surgery. However, the lack of age difference in recovery of thermal hypersensitivity after surgery suggests that sensitization of C-fiber input has a similar time course of resolution of pain over the ages studied in this model. The neural bases for these developmental differences are under study and may lead to a better understanding of pain during development and altered approaches to treatment of postoperative pain in neonates and infants.

Developmental and Regional Differences in Nitric Oxide Synthase Activity and Blood Flow in the Sheep Brain

Nitric oxide synthase (NOS) participates in the regulation of cerebral blood flow and neurotransmitter release and as a second messenger of glutamatergic and cholinergic systems. Developmental differences in NOS activity have been described in the rat, but not in a species with longer gestation and a larger, lobulated brain at birth. We assayed NOS activity by conversion of [14C]L-arginine to [14C]L-citrulline in 50-mg tissue samples from eight brain regions in sheep at 70, 92, 110, and 135 days gestation (term = 145 days); newborns (<7 days); and adults to test the hypothesis that NOS activity in the brain is developmentally regulated from midgestation through adulthood and matures along the neuroaxis in parallel with the known development of cerebral blood flow and neuronal activity. Three patterns of maturation of NOS activity were evident: increasing to or exceeding adult levels before 70 days gestation in the thalamus, cerebellum, and medulla; increasing to adult levels between 70 and 92 days in the hippocampus; and increasing to adult levels after 92 days in the cortex and caudate. Additionally, there were regional differences in cortical NOS activity: at 70 and 92 days of gestation, frontal cortex NOS activity was greater than parietal or occipital activity, and at 135 days gestation and in the newborn and adult, cortical and caudate activity exceeded that in most of the more caudal regions. The up to fourfold increase in regional cortical NOS activity between 92 and 135 days gestation was associated with twofold increases in cerebral blood flow and oxygen consumption during this period. Inhibition of NOS activity with administration of 60 mg/kg of NG-nitro-L-arginine methylester (L-NAME) resulted in 27% and 25% reductions in cerebral blood flow at 93 and 133 days gestation. While the associated increases in NOS activity with increases in CBF and CMRO2 do not appear causative, at various points in gestation the development of NOS activity may participate in the development of mature patterns of cerebral blood flow regulation in parallel with development of synaptic and electrical activity.