Fabrication of polylactic acid/paclitaxel nano fibers by electrospinning for cancer therapeutics.

Polylactic acid (PLA) is a thermoplastic and biodegradable polyester, largely derived from renewable resources such as corn starch, cassava starch and sugarcane. However, PLA is only soluble in a narrow range of solvents such as tetrahydrofuran, dioxane, chlorinated solvents and heated benzene. The limited choices of solvent for PLA dissolution have imposed significant challenges in the development of specifically engineered PLA nanofibers with electrospinning techniques. Generally, the electrospun polymeric materials have been rendered with unique properties such as high porosity and complex geometry while maintaining its biodegradability and biocompatibility for emerging biomedical applications. In this study, a new anticancer drug delivery system composed of PLA nanofibers with encapsulated paclitaxel was developed by the electrospinning of the respective nanofibers on top of a spin-coated thin film with the same chemical compositions. Our unique approach is meant for promoting strong bonding between PLA-based nanofibers and their respective films in order to improve the prolonged release properties and composite film stability within a fluctuative physiochemical environment during cell culture. PLA/paclitaxel nanofiber supported on respective polymeric films were probed by scanning electronic microscope, Fourier transform infrared spectrometer and water contact measurement for determining their surface morphologies, fibers' diameters, molecular vibrational modes, and wettability, respectively. Moreover, PLA/paclitaxel nanofibers supported on respective spin-coated films at different loadings of paclitaxel were evaluated for their abilities in killing human colorectal carcinoma cells (HCT-116). More importantly, MTT assays showed that regardless of the concentrations of paclitaxel, the growth of HCT-116 was effectively inhibited by the prolonged release of paclitaxel from PLA/paclitaxel nanofibers. An effective prolonged delivery system of paclitaxel based on PLA nanofiber-based film has demonstrated exciting potentials for emerging applications as implantable drug delivery patch in post-surgical cancer eradication.

Synthesis, characterization, and antibacterial activity of silver-doped silica nanocomposite particles.

Silver nanoparticles were adsorbed preferentially on silica surface to form composite particles using a reverse micelle process that stabilizes the silver particles by an anionic sodium bis(2-ethylhexyl) sulfosuccinate (AOT) surfactant in isooctane solvent together with the silica particles in which their surface being mediated by a cationic poly(allylamine hydrochloride) (PAH) polyelectrolyte. The heterogeneous adsorption was rendered by both electrostatic attraction and hydrophilic/hydrophobic interaction, and was carried out in multiple deposition cycles. The resulting nanocomposite particles were characterized by zeta-potential measurement, electron microscopy, X-ray diffractometry, field-emission electron spectroscopy for chemical analysis (ESCA), and inductively coupled plasma analysis, respectively. In addition, antibacterial activity of the composite particles was examined against Escherichia coli (E. coli) in aqueous environment.

Mechanisms underlying the cardioinhibitory and pressor responses elicited from the medullary neurons in the gigantocellular tegmental field of cats.

A stimulation of the gigantocellular tegmental field (FTG) in the medulla oblongata often increases systemic arterial blood pressure (SAP) and decreases heart rate (HR). We investigated if the cardioinhibitory/depressor areas, including the nucleus ambiguus (NA), the dorsal motor nucleus of vagus (DMV) and the caudal ventrolateral medulla (CVLM), underlied the functional expression of FTG neurons in regulating cardiovascular responses. In 73 chloralose-urethane anesthetized cats, the HR, SAP and vertebral nerve activity (VNA) were recorded. Neurons in the FTG, NA, DMV and CVLM were stimulated by microinjection of sodium glutamate (25 mM Glu, 70 nl). To study if the NA, DMV, and CVLM relayed the cardioinhibitory messages from the FTG, 24 mM kainic acid (KA, 100 nl) was used as an excitotoxic agent to lesion neurons in the NA, DMV or CVLM. We found that the cardioinhibition induced by FTG stimulation was significantly reduced by KA lesioning of the ipsilateral NA or DMV. Subsequently, a bilateral KA lesion of NA or DMV abolished the cardioinhibitory responses of FTG. Compared to the consequence of KA lesion of the DMV, only a smaller bradycardia was induced by FTG stimulation after KA lesion of the NA. The pressor response induced by Glu stimulation of the FTG was reduced by the KA lesion of the CVLM. Such an effect was dominant ipsilaterally. Our findings suggested that both NA and DMV mediated the cardioinhibitory responses of FTG. The pressor message from the FTG neurons might be partly working via a disinhibitory mechanism through the depressor neurons located in the CVLM.

The relationship between FTL and NA, DMV or CVLM in central cardiovascular control.

The aim of the present study was to examine the relationship between the lateral tegmental field (FTL), a cardioinhibitory area, with other cardioinhibitory areas, i.e., the ambiguus nucleus (NA) and the dorsal motor nucleus of vagus (DMV) and the caudal ventrolateral medulla (CVLM), a vasopressor inhibitory area. In 55 cats anesthetized with chloralose (40 mg/kg) and urethane (400 mg/kg), the cardiovascular responses of heart rate (HR), systemic arterial blood pressure (SAP) and vertebral nerve activity (VNA) were recorded. The FTL, NA, DMV and CVLM were identified first by stimulation (rectangular pulses in 80 Hz, 0.5 ms, 50-100 microA) and then confirmed by microinjection of sodium glutamate (Glu, 0.25M, 70 nl). In studying the influence of NA, DMV, or CVLM lesion on the Gluinduced responses in FTL, kainic acid (KA, 24 mM, 100 nl) was microinjected into the NA, DMV or CVLM. FTL stimulation produced an average decrease of HR by 55%. After KA lesioning of the ipsilateral NA or the DMV, the decreased HR induced by FTL was significantly diminished. After subsequent lesion of the contralateral DMV or NA, the bradycardia of FTL was abolished. The reduction of resting HR was more intense after lesioning the NA than DMV and with the left side more than that of the right side. These studies suggest that the cardioinhibitory responses of FTL are mediated through both NA and DMV with predominance of the former, while the hypotensive effect of FTL is mediated through CVLM. The precise pathway responsible for the FTL-induced bradycardia and hypotension is to be determined.

Measurement and simulation of light distribution in biological tissues.

Lateral light-distribution images of biologic tissues were used to study the tissues' optical characteristics. Monte Carlo simulation with the same conditions was performed to simulate the light distribution for comparison. Simulation results showed that the lateral light distribution was similar to the internal light distribution in biologic tissue. The direction of muscle fibers and the temperature both affect the near-field light distribution in tissue. The lateral view distribution can be both measured and simulated to study photon migration in tissue. It can also be used to estimate or verify the optical coefficients of tissue.

Use of extracorporeal membrane oxygenation to rescue a child with acute respiratory distress syndrome.

Acute respiratory distress syndrome (ARDS) carries a high mortality of about 60%. The results of conventional treatments for ARDS are poor. We report the use of extracorporeal membrane oxygenation (ECMO) to rescue a child with ARDS. The patient, a 7-year-old boy, underwent a Ross procedure and mitral valvuloplasty because of severe aortic and mild mitral regurgitation. ARDS due to massive transfusion and prolonged cardiopulmonary bypass developed in the early postoperative period. Hypoxemia persisted despite conventional treatments, including pressure-controlled ventilation and high-frequency ventilation. Finally, venovenous ECMO was used to rescue the patient. With ECMO support, gas exchange was well maintained with a lower ventilator setting, and ventilator-induced lung injuries were avoided. ECMO was used for 183 hours, at which point the boy was weaned without complications. His recovery was uneventful. At the latest follow-up, 6 months after the operation, he was in New York Heart Association function class I and had no complaints of lung disease. This case suggests that venovenous ECMO can be a rescue method for patients with ARDS that is refractory to conventional treatments.

A single minute lesion around the ventral respiratory group in medulla produces fatal apnea in cats.

In 35 adult cats anesthetized with intraperitoneal chloralose and urethane, the ventrolateral medulla was explored by microinjection of kainic acid (KA, 24 mM, 200 nl) with metal electrode-tubing or glass micropipette to determine regions which elicit persistent apnea. Persistent apnea is defined as: (1) In spontaneously breathing cats, termination of respiration over 3 min with a decrease of the mean systemic arterial pressure (MSAP) to 25 mm Hg. (2) In animals under artificial ventilation and paralyzed by gallamine, cessation of bilateral phrenic nerve (PNA) activities over 25 min. The apnea producing area was located dorsal to the rostral pole of the lateral reticular nucleus, ventromedial to the ambiguous nucleus and immediately caudal to the retrofacial nucleus. Functionally, this region includes the rostral part of the ventral respiratory group (rVRG) encompassing the pre-BOtzinger area. We define this region as the VRG apnea producing area (VRG-Apa). Fatal apneusis was observed under following conditions: (1) Persistent apnea was produced after a single KA microinjection in one side of the VRG-Apa (5 animals). Microinjection of sodium glutamate (0.25 M, 70-200 nl) in the same area produced only brief apnea, while microinjection of kynurenic acid (0.1 M, 200 nl) showed little effect on the respiration but slightly increased the SAP. (2) Positioning an electrode nearby but not in the VRG-Apa with or without KA injection did not produce apnea. But when a second electrode insertion to the opposite VRG-Apa immediately produced persistent apnea even without KA injection (6 animals). (3) Midsagittal division of the medulla 0-5 mm rostral to the obex produced persistent silence of PNA on both sides in artificial ventilated animals (7 animals), while similar division 0-5 mm caudal to the obex (4 animals) produced a brief but reversible quiescence of PNA. In conclusion, findings of the present study support the existence of a restricted region of VRG-Apa. VRG-Apa on both sides are closely connected, and integrity of both VRG-Apa is essential for normal respiration.

Vagal innervation of the gastrointestinal tract arises from dorsal motor nucleus while that of the heart largely from nucleus ambiguus in the cat.

The origin of medullary cells that form the cardiac vagal branch and the vagal branches in the lower thorax innervating the gastrointestinal (GI) tract was studied using horseradish peroxidase (HRP), a retrograde transport tracer in the cat. The distributions of parasympathetic postganglionic neurons of the heart were studied with acetylcholinesterase histochemistry. Intracardiac ganglionic neurons were found mainly in the connective tissue surrounding the base of the pulmonary arteries and in an area in and dorsal to the interatrial septum. Following injection of HRP into the subepicardum where most of the cardiac postganglionic neurons reside, 91% of the labelled neurons were found bilaterally distributed in the nucleus ambiguus (NA). A small population of labelled neurons was found in the dorsal motor nucleus of the vagus (DMV) and an intermediate zone (IZ) between the two nuclei. When HRP was injected into the left or right cardiopulmonary vagus branch, labelled neurons were found exclusively in the ipsilateral NA, DMV and IZ with a predominance in the NA. In the thorax, after they course around the heart, the left and right thoracic vagus nerves divides into a left and a right branch, respectively. The left branch of the left thoracic vagus joins the left branch of the right thoracic vagus to form the anterior vagus nerve at 3 cm above the diaphragm. The right branch of the right thoracic vagus nerve joins the right branch of the left thoracic vagus to form the posterior vagus nerve. After application of HRP into the right or the left branch of the left thoracic vagus, HRP labelled cells were found in the left DMV. Similarly, after application of HRP into the left or the right branch of the right thoracic vagus, labelled cells were found in the right DMV. On the other hand, when HRP was injected into the anterior vagus, labelled neurons were found bilaterally in the DMV. This suggests that all rostral branches of the thoracic vagus have their origin in the ipsilateral DMV, and intermixing occurs only at the caudal level near the diaphragm. Findings of the present experiments suggest that parasympathetic preganglionic neurons innervating the GI tract are located exclusively in the DMV while those of the heart are located mainly in the NA. Within the DMV, GI vagal neurons were found medially from the level 0-2.5 mm rostral to the obex. In contrast, cardiac vagal neurons were found in the lateral edge of the DMV at the level 0-1 mm rostral to the obex.

The depressor caudal ventrolateral medulla: its correlation with the pressor dorsomedial and ventrolateral medulla and the depressor paramedian reticular nucleus.

The functional correlation of the depressor caudal ventrolateral medulla (CVLM) with the two pressor regions, i.e. rostral ventrolateral medulla (RVLM) and dorsomedial medulla (DM), and with another inhibitory region, i.e. the paramedian reticular nucleus (PRN), were studied in cats anesthetized intraperitoneally with chloralose (40 mg/kg) and urethane (400 mg/kg). Systemic arterial pressure (SAP), heart rate (HR) and the sympathetic vertebral nerve activity (VNA) were recorded. The correct location of CVLM, RVLM or DM was determined by their specific responses, i.e. decreases of SAP, HR and VNA, for CVLM increases of these parameters for RVLM and DM, elicited first by electrical stimulation (80 Hz, 0.5 ms, 50-100 microA) then followed by microinjection of glutamate (Glu, 0.25 M, 70 nl). The depressor action of PRN was produced by electrical stimulation only. It was found that the depressor responses caused by the CVLM stimulation were greatly reduced 2 h after lesioning either the RVLM or DM by microinjection of kainic acid (KA, 24 mM, 200 nl) ipsilateral to the side of CVLM stimulation. The CVLM responses were further reduced after the remaining side of RVLM or DM was lesioned. The reduction of the CVLM-depressor responses was more apparent after the RVLM than DM lesioning. Data suggest that the CVLM-depressor responses are mediated through inhibition of the sympathetic-pressor neurons in both RVLM and DM with predominance of the former. Lesioning the PRN by KA and/or combination with DC electrolytic lesion reduced the CVLM-induced depressor responses. In turn, lesioning the CVLM by KA reduced the PRN-induced depressor responses. The reduction in the later manipulation was more apparent in the PRN-depressor responses than the CVLM-depressor responses. Data suggest that part of the PRN depressor action is mediated through activation of the CVLM.

Enterovesical fistula: 10 years experience.

BACKGROUND: Enterovesical fistula is a relatively uncommon complication of pelvic malignancies, diverticulitis of the colon, postoperative irradiation or trauma. Early diagnosis of enterovesical fistula is difficult and its management, complicated. The main purpose of this study is to investigate the most appropriate diagnostic method and to discuss the choice among different surgical managements. METHODS: From 1986 to 1995, 30 patients with enterovesical fistula were diagnosed and treated at Veterans General Hospital-Taipei. The records were reviewed retrospectively with regard to symptoms, primary disease process, diagnostic studies, management, complications, mortality and follow-up. RESULTS: Recurrent urinary tract infection (UTI) accounted for 73% and was the most common presenting symptom. Fecaluria (43%) and urine per rectum (40%) were another two common presenting symptoms. The major cause of these cases was malignancy (36%), followed by postoperative radiotherapy (17%) and iatrogenic injury (17%). Most of these cases were diagnosed by cystography (90%), barium enema (75%) or cystoscopy (69%). The rectum (52%) was the most common site involved, followed by the sigmoid colon (39%). The surgical management was individualized for each patient according to the general condition and the disease process. CONCLUSIONS: By combining the results of cystography, barium enema and cystoscopy, diagnosis of enterovesical fistula can be established in almost all cases. Single-stage operation for enterovesical fistula should be limited to those patients in good nutritional state and without severe inflammation, radiation injury, intestinal obstruction, other major medical problem, advanced malignancy or old age.

Effects of insulin on the cardiovascular integrating mechanisms of brain stem in cats.

In 65 cats anesthetized with alpha-chloralose and urethane, the effects of insulin on cardiovascular responses to stimulation of various structures in the brain stem were studied. The threshold dose of insulin injected intravenously that produced systemic hypoglycemia was 5-10 U/kg. Subthreshold hypoglycemic doses of insulin were used intracerebroventricularly (0.25 U/kg) or intracerebrally (2 mU in 200 nl). Sixty minutes after intravenous insulin, when serum glucose concentrations decreased from 158 to 43 mg/100 ml, pressor responses to stimulation of the periaqueductal gray of midbrain (PAG), locus coeruleus (LC), dorsal medulla (DM), ventrolateral medulla (VLM), and parvocellular reticular nucleus (PVC) decreased significantly. Depressor and bradycardiac response to stimulation of paramedian reticular nucleus or dorsal motor nucleus of vagus (DMV) decreased significantly as well. Thirty minutes after intracerebroventricular insulin, pressor responses of PAG, DM, and the bradycardiac response of DMV decreased significantly. Thirty minutes after intracerebral insulin, pressor responses and renal nerve activities of LC (but not PAG), VLM, DM, and PVC decreased significantly. A similar but faster onset (5 min) of depression of cardiovascular responses on stimulating the LC, VLM, DM, and PVC was observed in another six acutely midcollicular-decerebrate cats recovered from halothane anesthesia. These findings suggest that insulin directly inhibits the vasomotor structures of the brain stem and decreases the pressor responses to stimulation.

Long term and large-scale cultivation of human hepatoma Hep G2 cells in hollow fiber bioreactor. Cultivation of human hepatoma Hep G2 in hollow fiber bioreactor.

Long-term and large scale cultivation of an anchorage-dependent cell line using an industrial scale hollow fiber perfusion bioreactor is described. Hep G2 cells (a human hepatoma cell line) were cultivated in an Acusyst-P (Endotronic) with a total fiber surface area of 7.2 m2 6 x 1.2m2) to produce Hep G2 crude conditioned medium (CCM). Pretreatment of the cellulose acetate hollow fibers with collagen enhances the attachment of the anchorage-dependent cells. We have succeeded in growing the Hep G2 cells in an antibiotics- and serum-free IMDM medium, supplemented with 50 micrograms/ml of Hep G2 CCM protein at inoculation. The Hep G2 cells replicate and secrete CCM protein in quantities comparable to those produced in DMEM containing 10% fetal calf serum (FCS). The highest CCM protein productivity during the 80-day cultivation was 1.1 g/day with a total of 30 g of protein accumulated. Hep G2 CCM (20-40 micrograms protein/ml) was comparable to or even better than 10% FCS in supporting the growth of Molt-4 (a human T leukemia cell line) and FO (a mouse myeloma cell line) cells in vitro. The availability of this large amount of Hep G2 CCM will aid the further purification and characterization of growth factor(s) which could be used as serum substituents.

Interspecies differences in stereoselective protein binding and clearance of MK-571.

(+)-3-(((3-(2-(7-chloro-2-quinolinyl)ethenyl)phenyl)((3-(dimethylamino)- 3-oxopropyl)thio)methyl)thio)propanoic acid (MK-571), is a potent and specific antagonist of leukotriene D4 action in vitro and in vivo. The compound, which is being developed for the treatment of asthma, contains a chiral center at the methine carbon of the dithio side chain and exists in two forms. The binding of MK-571 enantiomers to plasma protein was extensive (greater than 99.5%), stereoselective, and species dependent. The R-(-)-enantiomer was bound to rat plasma to a greater extent than the S-(+)-enantiomer, while in dog and monkey plasma the reverse was the case. The elimination clearance of the enantiomers was inversely related to the stereoselective plasma protein binding, that with the greater unbound fraction being cleared more rapidly. Thus, the pharmacologically more active S-(+)-enantiomer was cleared 3.7 times more rapidly than its antipode in rats following iv administration of the racemate (10 mg/kg), whereas in dogs and monkeys the R-(-)-enantiomer was cleared more rapidly. Kinetic analysis of the data revealed that the intrinsic clearances of the unbound enantiomers were similar within species, suggesting that stereoselectivity in elimination is not attributable to differences in metabolism and biliary excretion. Bioavailabilities of the S-(+)- and R-(-)-enantiomers in the rat were similar (75% and 71%, respectively) suggesting that MK-571 was not stereoselectively absorbed in that species.

[Large-scale production of monoclonal antibodies using hollow fiber bioreactor].

Development of large-scale monoclonal antibody production using industrial scale hollow fiber bioreactor is described. Hybridoma cell lines H505AC and A306AA were cultivated in the hollow fiber perfusion bioreactor with a total fiber surface area of 7.2 m2 (6 x 1.2 m2) to produce anti-hepatitis B surface antigen (anti-HBsAg) monoclonal antibody IgM and anti-alpha-fetal protein (anti-AFP) monoclonal antibody IgG. The highest anti-HBsAg IgM productivity obtained during 49 days cultivation of H505AC was 0.135 g/day and the total accumulated IgM was 1.90 g. Results from the cultivation of A306AA in the hollow fiber module for 23 days showed the highest anti-AFP IgG productivity and the total accumulated IgG were 0.672 g/day and 7.21 g, respectively.

Persistence of renal nerve and spinal reflex activities and visceral motility after asphyxial anoxia.

Phrenic and renal nerves activities, spinal reflex (knee jerk and the evoked monosynaptic reflex response from the 5th lumbar ventral root), motility of the stomach, duodenum, jejunum and colon, and motility of urinary bladder during and after the lethal asphyxia were studied in cats under intraperitoneal urethane (400 mg/kg) and chloralose (40 mg/kg) anesthesia. Asphyxial anoxia produced one or two peaks of elevation of systemic arterial pressure (SAP) followed by a progressive decrease of SAP and narrowing of pulse pressure and eventual complete cardiovascular arrest. The phrenic nerve activity increased markedly during the final asphyxial SAP elevation; it then decreased along with the decline of SAP and ceased permanently when the SAP reached about 50 mmHg. The renal nerve activity increased along with each phase of pressure elevation. During the terminal stage of progressive SAP fall, the renal nerve activity fluctuated up and down 2-3 times before the SAP dropped to zero, and the activity persisted for an average of 202 sec thereafter. Data suggest that the motoneurons of the phrenic nerve in the cervical spinal cord and medulla oblongata are more vulnerable to asphyxia than the motoneurons responsible for the spinal reflex in the pathway from medulla, the intermediolateral column of the spinal cord and the celiac ganglion. During anoxia, the lumbar spinal reflex showed an initial inhibition then potentiation afterward. Even after the SAP had reached zero, the spinal reflex could still be elicited for one to a few minutes. This suggests that the spinal cord can function sometime after complete cardiac arrest subsequent to asphyxia. The motility of the stomach, duodenum, jejunum, colon and urinary bladder usually decreased during the period of pressure elevation. However, the activity was enhanced during the terminal stage of pressure decline. The enhancement of the colon and bladder motility, spasmodic in nature, was particularly prominent, more so than the others. After complete cardiovascular arrest, the motility of the above viscera became temporary quiescent but about 10 minutes later, it resumed activity again with an enhancement of action in some of the animals. The motility, persisted from few minutes to over six hours. These phenomena indicate that despite a complete deprivation of circulatory and respiratory supports, or death of the animal, the viscera can still contract for a certain period of time.

Oxidative bioconversion of toluene to 1,3-butadiene-1,4-dicarboxylic acid (cis,cis-muconic acid).

Pseudomonas sp. strains, isolated from soil, utilized toluene as their sole carbon source through ameta cleavage pathway. Strains metabolizing toluene through anortho cleavage pathway were selected from the wild typemeta strain. Theortho pathway strains were subjected to chemostat selection to obtain a fast-growing strain with doubling time reduced from 14 to 1.2 h. Benzoale and antibiotics enrichment selection procedures were utilized to select a blocked mutant. The blocked mutant grew on acetate as its sole carbon source and oxidatively converted toluene tocis, cis-muconic acid. Double-blocked and muconate-permeable mutants were also selected to reduce reversion frequency and to enhance muconic acid production. In shake-flask experiments, muconic acid at 3.5 g/l was obtained after 2 days of fermentation. In a 14 l fermenter, muconic acid was produced at 45 g/l in 4 days of controlled fed-batch fermentation. The oxidative bioconversion process was also demonstrated in a 1500 l fermenter.

Predominance of vagal bradycardia mechanism in the brain stem of turtles.

Cardiovascular parameters of spontaneously breathing pond turtles (Cyclemys flavomarginata) anaesthetized with chloralose (4 mg 100 g-1) and urethane (40 mg 100 g-1), were examined during exploratory electrical stimulation of the brain stem. Turtles exhibited a low mean systemic arterial blood pressure (MSAP, average 25 mmHg) and slow heart rate (average 24 beats min-1). Upon stimulation, pressor (sympathetic), depressor (sympathetic inhibition), bradycardia and hypotensive (vagal) responses were elicited from regions of the brain stem extending from the hypothalamus to the medulla, principally in the medial region. The pressor response appeared after a longer latency than did the bradycardia and hypotensive responses. It developed rather slowly, and rarely attained a magnitude double its resting value. In contrast, stimulation of many points in the brain stem produced marked slowing or even cessation of the heart beat, and thus resulted in an immediate fall of the blood pressure even to zero. This cardio-inhibitory response depended on the integrity of the vagus nerves and was particularly marked upon stimulation in the caudal medulla, the areas of the ambiguus, solitary and dorsomotor nuclei of the vagus and the midline structures. When such an area was stimulated continuously the heart stopped beating throughout the stimulation. The longest period of cardiac arrest before the appearance of escape was 35 min. With continuous stimulation of the peripheral end of the cut vagus, the earliest escape beat occurred even later (65 min). Epinephrine given intravenously produced an increase of MSAP and force of cardiac contraction, although the slope of pressor rise was shallow. Reflex bradycardia, however, was not observed. These experiments show that a very prominent vagal bradycardia can be evoked from the turtle brain stem, which may contribute to its well-known capacity for tolerating anoxia.