Is there a rational basis for cannabinoids research and development in ocular pain therapy? A systematic review of preclinical evidence.

BACKGROUND: Purpose of the present systematic review is to investigate preclinical evidence in favor of the working hypothesis of efficacy of cannabinoids in ocular pain treatment. METHODS: Literature search includes the most relevant repositories for medical scientific literature from inception until November, 24 2021. Data collection and selection of retrieved records adhere to PRISMA criteria. RESULTS: In agreement with a priori established protocol the search retrieved 2471 records leaving 479 results after duplicates removal. Eleven records result from title and abstract screening to meet the inclusion criteria; only 4 results are eligible for inclusion in the qualitative synthesis impeding meta-analysis. The qualitative analysis highlights the antinociceptive and anti-inflammatory efficacy of Δ8-tetrahydrocannabinol, cannabidiol and its derivative HU-308 and of new racemic CB1 allosteric ligand GAT211 and its enantiomers GAT228 and GAT229. Moreover, CB2R agonists RO6871304 and RO6871085 and CB2R ligand HU910 provide evidence of anti-inflammatory efficacy. CB2 agonist HU308 reduces of 241% uveitis-induced leukocyte adhesion and changes lipidome profile. Methodological and design issues raise concern of risk of bias and the amount of studies is too small for generalization. Furthermore, the ocular pain model used can resemble only inflammatory but not neuropathic pain. CONCLUSIONS: The role of the endocannabinoid system in ocular pain is underinvestigated, since only two studies assessing the effects of cannabinoid receptors modulators on pain behavior and other two on pain-related inflammatory processes are found. Preclinical studies investigating the efficacy of cannabinoids in ocular inflammatory and neuropathic pain models are needed to pave the way for clinical translation.

Influence of the polymorphism of the DUSP14 gene on the expression of immune-related genes and development of pulmonary tuberculosis.

Recently, a genome-wide screening identified a functional single-nucleotide polymorphism in dual-specificity phosphatase 14 gene (DUSP14), which was associated with pulmonary tuberculosis (TB) in a West African study. DUSP14 regulates T-cell proliferation and cytokine production in a negative way via dephosphorylation and inactivation of key signaling molecules. The aim of this study is to further explore the possible significance of the DUSP14 polymorphism. Total RNA was extracted from the whole blood of 109 healthcare workers (HCWs) in Vietnam and subjected to quantitative reverse-transcription PCR for DUSP14 and 20 immune-related genes. DUSP14 rs1051838 was genotyped in 502 new pulmonary TB patients and 506 healthy controls. Among disease-free individuals (HCWs), T-helper type-1 (Th1)-related genes, interferon-gamma receptor 2 (IFNGR2) and signal transducer and activator of transcription-1 (STAT1) mRNA levels significantly increased as the number of A alleles of rs1051838 increased, whereas the DUSP14 mRNA level tended to decrease. The AA genotype was associated with protection against active TB in younger patients (⩽45 years old, OR=0.63, 95% CI 0.44-0.90). Our results suggest that a low-expression genotype of DUSP14 accompanied by high transcript levels of Th1 immune-related genes may confer protection against early TB development.

Rational Basis for the Use of Bergamot Essential Oil in Complementary Medicine to Treat Chronic Pain.

In complementary medicine, aromatherapy uses essential oils to improve agitation and aggression observed in dementia, mood, depression, anxiety and chronic pain. Preclinical research studies have reported that the essential oil obtained from bergamot (BEO) fruit (Citrus bergamia, Risso) modifies normal and pathological synaptic plasticity implicated, for instance, in nociceptive and neuropathic pain. Interestingly, recent results indicated that BEO modulates sensitive perception of pain in different models of nociceptive, inflammatory and neuropathic pain modulating endogenous systems. Thus, local administration of BEO inhibited the nociceptive behavioral effect induced by intraplantar injection of capsaicin or formalin in mice. Similar effects were observed with linalool and linalyl acetate, major volatile components of the phytocomplex, Pharmacological studies showed that the latter effects are reversed by local or systemic pretreatment with the opioid antagonist naloxone hydrochloride alike with naloxone methiodide, high affinity peripheral µ-opioid receptor antagonist. These results and the synergistic effect observed following systemic or intrathecal injection of an inactive dose of morphine with BEO or linalool indicated an activation of peripheral opioid system. Recently, in neuropathic pain models systemic or local administration of BEO or linalool induced antiallodynic effects. In particular, in partial sciatic nerve ligation (PSNL) model, intraplantar injection of the phytocomplex or linalool in the ipsilateral hindpaw, but not in the contralateral, reduced PSNL-induced extracellularsignal- regulated kinase (ERK) activation and mechanical allodynia. In neuropathic pain high doses of morphine are needed to reduce pain. Interestingly, combination of inactive doses of BEO or linalool with a low dose of morphine induced antiallodynic effects in mice. Peripheral cannabinoid and opioid systems appear to be involved in the antinociception produced by intraplantar injection of ß -caryophyllene, present in different essential oils including BEO. The data gathered so far indicate that the essential oil of bergamot is endowed with antinociceptive and antiallodynic effects and contribute to form the rational basis for rigorous testing of its efficacy in complementary medicine.

Involvement of peripheral cannabinoid and opioid receptors in ß-caryophyllene-induced antinociception.

BACKGROUND: ß-caryophyllene (BCP) is a common constitute of the essential oils of numerous spice, food plants and major component in Cannabis. The present study investigated the contribution of peripheral cannabinoid (CB) and opioid systems in the antinociception produced by intraplantar (i.pl.) injection of BCP. The interaction between peripheral BCP and morphine was also examined. METHODS: The antinociceptive effect of i.pl. BCP was assayed by the capsaicin tests in mice. Antagonists for CB and opioid receptors, and antisera against ß-endorphin were injected peripherally prior to i.pl. injection of BCP. Morphine in combination with BCP was injected subcutaneously or intrathecally. RESULTS: The i.pl. injection of BCP dose-dependently attenuated capsaicin-induced nociceptive response. The antinociceptive effect produced by BCP was prevented by pretreatment with AM630, a selective CB2 receptor antagonist, but not by AM251, a selective CB1 receptor antagonist. Pretreatment with naloxone, an opioid receptor antagonist, and ß-funaltrexamine, a selective µ-opioid receptor antagonist, reversed the antinociceptive effect of BCP. Pretreatment with naloxone methiodide, a peripherally acting antagonist for opioid receptors and antisera against ß-endorphin, resulted in a significant antagonizing effect on BCP-induced antinociception. Morphine-induced antinociception was increased by a low dose of BCP. The increased effect of morphine in combination with BCP was antagonized significantly by pretreatment with naloxone. CONCLUSIONS: The present results demonstrate that antinociception produced by i.pl. BCP is mediated by activation of CB2 receptors, which stimulates the local release from keratinocytes of the endogenous opioid ß-endorphin. The combined injection of morphine and BCP may be an alternative in treating chemogenic pain.

Enteric opportunistic parasitic infections among HIV seropositive patients in Kathmandu, Nepal.

BACKGROUND: Enteric opportunistic parasitic infections are the major source of diarrheal disease in developing countries mainly in Human Immunodeficiency virus (HIV) infected patients. OBJECTIVE: The study was to detect enteric parasites causing diarrhea and their association with immune status in HIV-seropositive patients. METHODS: The present study was conducted in Dirgh-Jeevan Health Care Research Center and Tribhuvan University Teaching Hospital, Public Health Research Laboratory, Kathmandu, Nepal between June 2010 and May 2011 involving 146 Human Immunodeficiency virus (HIV) positive patients. Serostatus from these patients were detected by Enzyme Linked Immunosorbent assay. CD4+ T cell counts were done by flow cytometry. Stool was examined for enteric parasites by microscopy with special staining methods. RESULTS: A total of 146 HIV sero-positive patients with and without diarrhea age between 20 to 45 years were included in the study. Of the 146 patients, the protozoan parasitic infection was found in 30.13% (44/146). Out of 146 patients, 78 had diarrhea in which parasitic infection was 39 (50%) and 7.35% (5/68) protozoal parasites positive cases did not have diarrhea. A significant difference (p less than 0.05) was observed in the level of infection of intestinal protozoan between the HIV seropositive with diarrhea and HIV-seropositive without diarrhea. Out of 43 patients whose CD4+ T cells were less than 200/µl, 29 (67.4%) had opportunistic parasitic infection whereas out of 103 patients whose CD4+ T cells were =200/mcl, only 15 (14.56%) had opportunistic parasitic infection (P less than 0.05). CONCLUSION: Enteric opportunistic parasitic infections were detected in 30.1% among HIV-seropositive patients and low CD4+ T count indicated high enteric opportunistic infection. Early detection of enteric parasitic infections will help in the management and to improve the quality of life for HIV-infected individuals.

HLA-A, -B, -C, -DRB1 and -DQB1 alleles and haplotypes in the Kinh population in Vietnam.

Allele and haplotype frequencies of the human leukocyte antigens (HLA) were studied in the Kinh Vietnamese population. We analyzed 170 unrelated healthy individuals. DNA-based HLA typing was performed using a microsphere-based array genotyping platform with sequence-specific oligonucleotide probes to distinguish HLA-A, -B, -C, -DRB1 and -DQB1 alleles. A total of 21 HLA-A, 37 HLA-B, 18 HLA-C, 25 HLA-DRB1, and 14 HLA-DQB1 alleles were identified. HLA-A*1101, A*2402, A*3303, B*1502, B*4601, Cw*0102, Cw*0702, Cw*0801, DRB1*1202, DQB1*0301, DQB1*0303, and DQB1*0501 were found with frequencies higher than 10%. Two representative haplotypes bearing two to five HLA loci were A*1101-B*1502 and A*3303-B*5801 for HLA-A-B; Cw*0801-B*1502 and Cw*0102-B*4601 for HLA-C-B; B*1502-DRB1*1202 and B*4601-DRB1*0901 for HLA-B-DRB1; DRB1*1202-DQB1*0301 and DRB1*0901-DQB1*0303 for HLA-DRB1-DQB1; A*1101-Cw*0801-B*1502 and A*3303-Cw*0302-B*5801 for HLA-A-C-B; A*1101-B*1502-DRB1*1202 and A*2901-B*0705-DRB1*1001 for HLA-A-B-DRB1, A*1101-Cw*0801-B*1502-DRB1*1202-DQB1*0301 and A*2901-Cw*1505-B*0705-DRB1*1001-DQB1*0501 for HLA-A-C-B-DRB1-DQB1. Allele distribution and haplotype analysis demonstrated that the Vietnamese population shares HLA patterns with southern Chinese, Thai, Javanese and Micronesians, while it also retains unique characteristics.

Pulmonary Mycobacterium avium complex infection associated with the IVS8-T5 allele of the CFTR gene.

BACKGROUND: The T5 allele in intron 8 (IVS8) on specific haplotype backgrounds (e.g., long TG repeats) causes abnormal splicing in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, and is also known to be associated with chronic airway diseases. OBJECTIVE: To investigate the role of CFTR variations for susceptibility to pulmonary Mycobacterium avium complex (MAC) infection. PARTICIPANTS: Three hundred patients with pulmonary MAC infection (72 males, 228 females; mean age at onset 61.6 + or - 12.4 years) took part in this study. Diagnosis of MAC infection was based on American Thoracic Society criteria. Clinical profiles were collected and blood samples were genotyped for TG repeats, poly-T and M470V polymorphisms. RESULTS: We found significantly higher T5 frequency in MAC patients than in healthy controls from our own study (0.035 and 0.005, respectively, P = 0.023) and other reports. Homozygote for the T5 allele was found in two MAC patients. All T5 alleles were associated with longer TG repeats, the TG12 or TG13 allele. Seventeen of the 21 T5 alleles appeared to be associated with the V470 allele. Other polymorphisms did not show any significant differences in frequency. CONCLUSIONS: These findings suggest that the IVS8 5T allele might be involved in susceptibility to pulmonary MAC infection.

A survey of tuberculosis prevalence in Hanoi, Vietnam.

OBJECTIVE: To assess the prevalence of tuberculosis (TB) in Hanoi, Vietnam, in 2003/2004. METHODS: A random selection was carried out involving 11624 subjects from 20 communes within the city. RESULTS: On chest X-ray examination, 317 subjects (2.73%) showed abnormal lung opacity, of which 17 were sputum smear-positive, two concentrated smear-positive and three culture-positive, all with active TB. The prevalence of sputum smear-positive pulmonary TB was 146 per 100000 in persons aged >or=15 years (95%CI 65-228). CONCLUSION: This is the first large-scale assessment of the prevalence of TB in Hanoi. The prevalence rate was higher than expected, suggesting that a significant number of patients with active TB, particularly females, remain undiagnosed, thus representing a continuing potential source of transmission in the community.

Differential effects of yohimbine, naloxone and 8-OH-DPAT on ejaculatory response in male dogs.

The aim of this study was to compare the effects of the alpha(2)-adrenergic-receptor antagonist yohimbine, the 5-HT(lA)-receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and the opioid-receptor antagonist naloxone (all of which have been shown to stimulate male sexual arousal/motivation in rats) on sexual responses in male dogs. Sexual responses (i.e., ejaculation, penile erection and pelvic thrusting behavior) were elicited by manual penile stimulation. Systemic administration of yohimbine (0.03-1.0 mg/kg) produced a biphasic dose response curve for the amount of ejaculated semen collected during genital stimulation (for 5 min), whereas 8-OH-DPAT (0.03-0.3 mg/kg) dose-dependently decreased the amount of ejaculated semen. Thus, yohimbine increased the amount of ejaculated semen at lower doses (0.03-0.3 mg/kg), but decreased it at the highest dose (1.0 mg/kg). The highest dose of yohimbine (1.0 mg/kg) and 8-OH-DPAT (0.3 mg/kg) also produced a significant delay of onset in both ejaculation and penile erection latency (time from starting the stimulation to the first ejaculation and full erection), and a decrease in the incidence of pelvic thrusting behavior. In contrast, administration of naloxone (0.03-1.0 mg/kg) did not affect the sexual responses elicited by genital stimulation. These results indicate that yohimbine and 8-OH-DPAT, but not naloxone, affect sexual responses, particularly ejaculation, and that the drugs which stimulate the mechanisms regulating sexual arousal/motivation in male rats do not show identical effects for sexual function in male dogs. The present findings also confirm our previous observations that the ejaculatory capacity in dogs can be stimulated by lower doses of yohimbine, as evidenced by an increase in the amount of ejaculated semen.

Involvement of adenosine A1 receptors in forced walking stress-induced analgesia in mice.

We investigated the involvement of adenosine receptors on forced walking stress-induced analgesia using a formalin-induced paw-licking test in male mice. Exposure to forced walking stress for 6 h showed stress-induced analgesia in the second phase (10-30 min), but not in the first phase (0-10 min). In the second phase, forced walking stress-induced analgesia was blocked by theophylline, a nonselective adenosine-receptor antagonist and DPCPX, an adenosine A1-receptor antagonist, but not ZM 241385, an adenosine A2A-receptor antagonist. These findings suggest that adenosine A1 receptors are involved in the analgesic mechanism activated by the forced walking stress.

Differential antinociceptive effects induced by intrathecally administered endomorphin-1 and endomorphin-2 in the mouse.

Two highly selective mu-opioid receptor agonists, endomorphin-1 and endomorphin-2, have been identified and postulated to be endogenous ligands for mu-opioid receptors. Intrathecal (i.t.) administration of endomorphin-1 and endomorphin-2 at doses from 0.039 to 5 nmol dose-dependently produced antinociception with the paw-withdrawal test. The paw-withdrawal inhibition rapidly reached its peak at 1 min, rapidly declined and returned to the pre-injection levels in 20 min. The inhibition of the paw-withdrawal responses to endomorphin-1 and endomorphin-2 at a dose of 5 nmol observed at 1 and 5 min after injection was blocked by pretreatment with a non-selective opioid receptor antagonist naloxone (1 mg/kg, s.c.). The antinociceptive effect of endomorphin-2 was more sensitive to the mu (1)-opioid receptor antagonist, naloxonazine than that of endomorphin-1. The endomorphin-2-induced paw-withdrawal inhibition at both 1 and 5 min after injection was blocked by pretreatment with kappa-opioid receptor antagonist nor-binaltorphimine (10 mg/kg, s.c.) or the delta(2)-opioid receptor antagonist naltriben (0.6 mg/kg, s.c.) but not the delta(1)-opioid receptor antagonist 7-benzylidine naltrexone (BNTX) (0.6 mg/kg s.c.). In contrast, the paw-withdrawal inhibition induced by endomorphin-1 observed at both 1 and 5 min after injection was not blocked by naloxonazine (35 mg/kg, s.c.), nor-binaltorphimine (10 mg/kg, s.c.), naltriben (0.6 mg/kg, s.c.) or BNTX (0.6 mg/kg s.c.). The endomorphin-2-induced paw-withdrawal inhibition was blocked by the pretreatment with an antiserum against dynorphin A-(1-17) or [Met(5)]enkephalin, but not by antiserum against dynorphin B-(1-13). Pretreatment with these antisera did not affect the endomorphin-1-induced paw-withdrawal inhibition. Our results indicate that endomorphin-2 given i.t. produces its antinociceptive effects via the stimulation of mu (1)-opioid receptors (naloxonazine-sensitive site) in the spinal cord. The antinociception induced by endomophin-2 contains additional components, which are mediated by the release of dynorphin A-(1-17) and [Met(5)]enkephalin which subsequently act on kappa-opioid receptors and delta(2)-opioid receptors to produce antinociception.

Alpha 2-adrenoceptor antagonists: effects on ejaculation, penile erection and pelvic thrusting behavior in dogs.

We previously reported that systemic administration of yohimbine, an alpha2-adrenoceptor antagonist, exerts a biphasic effect (stimulating and suppressing) on ejaculation in dogs, when this function is analyzed using the amount of ejaculated semen in response to genital stimulation. To clarify the effect of alpha2-adrenoceptor blockade on male sexual function, we investigated the effects of four selective alpha2-adrenoceptor antagonists, rauwolscine, idazoxan, RX821002 and mydaglizole, on sexual responses (ejaculation, penile erection and pelvic thrusting behavior) elicited by manual penile stimulation in dogs. Rauwolscine (intraperitoneal, 30 min before the testing) caused a biphasic effect on ejaculation; the amount of ejaculated semen produced by the stimulation was significantly increased by the lower doses (0.1 and 0.3 mg/kg), whereas it was decreased by the higher doses (1.0 and 2.0 mg/kg). The higher doses of rauwolscine also markedly inhibited both penile erection and pelvic thrusting behavior. Idazoxan and RX821002, at doses of 0.1 and 0.3 mg/kg, caused a significant increase in the amount of ejaculated semen without affecting other sexual functions. RX821002 (2.0 mg/kg), but not idazoxan (2.0 mg/kg), moderately inhibited both penile erection and pelvic thrusting behavior. Mydaglizole, a peripherally acting alpha2-adrenoceptor antagonist, did not affect the sexual responses at any doses (0.1-4.0 mg/kg). In the ejaculatory declining test, all alpha2-adrenoceptor antagonists (0.1 mg/kg), except for mydaglizole, completely prevented the decrease in ejaculatory capacity produced by antecedent ejaculation. These results indicate that, though the range of the effective dose is narrow, the alpha2-adrenoceptor antagonists that can block the central alpha2-adrenoceptors have the stimulatory effects on ejaculatory function. The difference of the sexual effects may be based on the action except for the alpha2-adrenoceptor blockade.

Rho activation in excitatory agonist-stimulated vascular smooth muscle.

Small GTPase Rho and its downstream effector, Rho kinase, have been implicated in agonist-stimulated Ca(2+) sensitization of 20-kDa myosin light chain (MLC(20)) phosphorylation and contraction in smooth muscle. In the present study we demonstrated for the first time that excitatory receptor agonists induce increases in amounts of an active GTP-bound form of RhoA, GTP-RhoA, in rabbit aortic smooth muscle. Using a pull-down assay with a recombinant RhoA-binding protein, Rhotekin, we found that a thromboxane A(2) mimetic, U-46619, which induced a sustained contractile response, induced a sustained rise in the amount of GTP-RhoA in a dose-dependent manner with an EC(50) value similar to that for the contractile response. U-46619-induced RhoA activation was thromboxane A(2) receptor-mediated and reversible. Other agonists including norepinephrine, serotonin, histamine, and endothelin-1 (ET-1) also stimulated RhoA, albeit to lesser extents than U-46619. In contrast, ANG II and phorbol 12,13-dibutyrate failed to increase GTP-RhoA. The tyrosine kinase inhibitor genistein substantially inhibited RhoA activation by these agonists, except for ET-1. Thus excitatory agonists induce Rho activation in an agonist-specific manner, which is thought to contribute to stimulation of MLC(20) phosphorylation Ca(2+) sensitivity.

Effective human herpesvirus 8 infection of human umbilical vein endothelial cells by cell-mediated transmission.

Cell-free transmission of human herpesvirus 8 (HHV-8) to human cells in vitro has been reported to be difficult, if not impossible. The present experiments were conducted with the idea that cell-cell contact may produce much more effective transmission, so-called cell-mediated transmission. Primary human umbilical vein endothelial cells (HUVECs) were cocultured with an HHV-8-infected lymphoma cell line, BCBL-1 cells. When a ratio of 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated BCBL-1 cells to HUVECs of 10:1 was used, more than 20% of HUVECs were found to express the HHV-8 latency-associated nuclear antigen (LANA) 48 h after the start of coculturing; this value increased to more than 30% after 72 h. HHV-8-encoded ORF26, K8, K8.1, K10, K11, ORF59, and ORF65 proteins were not detected in these HHV-8-infected HUVECs until 72 h. The HHV-8 antigens were not observed in HUVECs cocultured with TPA-treated BCBL-1 cells separated by a membrane. Thirty days after removal of the BCBL-1 cells from the cell-mediated transmission experiment, the HUVECs still expressed LANA and the HHV-8 genome was detected by PCR in these cells. Moreover, the ORF59 protein, a DNA replication-associated protein of HHV-8, was expressed in such HUVECs in the presence of TPA stimulation. These results indicated a far more effective transmission mechanism, cell-cell contact, suggesting the possibility that such a mechanism works in vivo.

Subtype-specific, differential activities of the EDG family receptors for sphingosine-1-phosphate, a novel lysophospholipid mediator.

The lysosphingolipid sphingosine-1-phosphate (S1P) and the structurally related lipid lysophosphatidic acid (LPA) elicit a wide spectrum of biological responses in a variety of cell types, including mitogenesis, cell-shape changes, migration and contraction. Recent studies have unveiled the existence of the G protein-coupled heptahelical receptor subfamily for the biologically active lysophospholipids, which consists of the two receptor subgroups specific for S1P and LPA, respectively. The S1P receptor subgroup comprises four members, i.e. EDG-1, EDG-3, EDG-5/AGR16 and EDG-6, with considerable amino acid similarity among them. The S1P receptor subtypes are coupled to different heterotrimeric G proteins, leading to the activation of a unique set of multiple intracellular signaling pathways. The expression of transcripts of the S1P receptor subtypes is wide-spread, except for EDG-6 which exhibits lymphoid tissue-specific expression. Plasma contains substantial concentrations of S1P as well as LPA. Activated platelets appear to be a major source of S1P and LPA in blood. In addition, accumulating evidence demonstrates that S1P and LPA are released from a variety of cell types in response to various extracellular stimuli. These observations demonstrate the existence of the novel signaling system comprising the lysosphingolipids and their cognate receptors, suggesting physiological and pathological roles.

Structure-antinociceptive activity studies of incarvillateine, a monoterpene alkaloid from Incarvillea sinensis.

Incarvillateine (1), a new monoterpene alkaloid carrying a characteristic cyclobutane ring, has been found to show significant antinociceptive activity in a formalin-induced pain model in mice. To investigate the correlation between its structure and antinociceptive activity, and especially to study whether a cyclobutane ring is necessary or not for expression of activity, we evaluated the antinociceptive activity of two constructive units of incarvillateine, such as a monoterpene unit (incarvilline, 3) and a phenylpropanoid unit (ferulic acid, 2) in the formalin test, and compared activity of the units with that of incarvillateine. Furthermore, in order to obtain more information about the structure-activity relationships, monoterpene alkaloid derivatives, such as incarvine C (5, a precursor of incarvillateine), incarvine A (4, an ester compound comprised of two monoterpene alkaloids and a monoterpene) and 3,3'-demethoxy-4,4'-dehydroxyincarvillateine (6, a synthetic new compound), were examined. The antinociceptive effect of 3,3'-demethoxy-4,4'-dehydroxyincarvillateine was equal to that of incarvillateine. Meanwhile, the other compounds exhibited no or weak activity. These results suggested that the cyclobutane moiety of incarvillateine plays an important role in expression of antinociceptive action.

Enzymatic determination of acetylcarnitine for diagnostic applications.

An enzymatic method was proposed for determination of acetylcarnitine (AcCar), even when carnitine (Car), non-acetylated form, co-exists. The method is consisted of four enzymatic reactions: First, AcCar is hydrolysed by acylcarnitine hydrolase to yield acetate; followed by the other three reactions coupled with three enzymes, respectively, acetate kinase, pyruvate kinase and lactate dehydrogenase; finally, the acetate formation causes a decrease in NADH. The amount of AcCar is then evaluated as the change in absorbance at 340 nm. The reagent composition of the reaction mixture was determined, and the characteristics of the method were investigated. The dilution test showed a good linearity over a wide range. The precision and accuracy tests produced satisfactory results. The co-existence of Car gave no effect on the measurement. The present method was found to be used easily, simply and rapidly for the selective determination of AcCar.

Ambient temperatures preferred by humans acclimated to heat given at a fixed daily time.

We investigated preferred ambient temperatures (T(pref)) of heat-acclimated humans to assess their behavioral thermoregulation. Seven male volunteers were exposed to an ambient temperature (T(a)) of 42 degrees C and relative humidity (RH) of 40% for 4 h (14:00-18:00 h)/day for 9-10 consecutive days. Rectal temperature (T(re)) was measured, and T(pref) was determined at two distinct times of day, 09:00-11:00 h (AM test) and 14:00-16:00 h (PM test), in both heat- and nonheat-acclimated (control) conditions. Heat acclimation significantly decreased T(re) only in the PM test. There was no difference in the T(pref) between the two tests in the control condition. However, T(pref) in the PM test was significantly lower than that of the AM test in the heat-acclimated condition. The findings suggest that repeated heat exposure in humans for 4 h at a fixed time daily alters the core temperature level and behavioral thermoregulatory function, particularly during the period when the subjects had previously been exposed to heat.