FLAVONOID CHEMISTRY AND SYSTEMATICS OF TITHONIA (COMPOSITAE)

Twenty-four flavonoid compounds, including chalcones and aurones, are reported from 13 taxa of Tithonia. Pedunculin (4′, 6, 7-trimethoxy, 5, 8-dihydroxyflavone), also is characterized. The distribution of these compounds is compared to the existing classification of the genus based primarily on morphological data. By comparison with the previously proposed phylogeny of the genus, the evolution of the flavonoid system may be elucidated. It is proposed that the ancestral stock of Tithonia had flavonols, di- and tri-methoxyflavones, and 6-methoxyflavones. From this ancestral stock, a number of trends have occured in derivative taxa. These trends are: 1) a maintenance of a diversity of compounds in several evolutionary lines, 2) a reduction in the number of methoxyl substitutions, and 3) the elaboration of 5-deoxyflavones and flavonols.     

Comparison of soluble and membrane polypeptides ofSynechocystis 6308 by two-dimensional electrophoresis

Two-dimensional electrophoresis was carried out on fractions of the cyanobacteriumSynechocystis 6308 (ATCC 27150). Phycobilisomes isolated fromSynechocystis in 0.75M potassium phosphate buffer, pH 6.8, (KPi) plus Triton X-100 showed 5 prominent polypeptides when examined by two-dimensional electrophoresis. Ultracentrifugation of cells broken in KPi lacking Triton yielded three fractions, a membrane-containing pellet, a green supernatant, and a less dense yellow supernatant. The three fractions yielded a total of 272 polypeptides visualized by silver staining of two-dimensional gels. Fourteen polypeptides were found only in the yellow fraction, 14 polypeptides were found only in the green fraction, and 16 polypeptides were found only in the membrane fraction; 23 polypeptides were found in all three fractions. The crude Triton-containing KPi extract contained 62, and a 50 mM HEPES extract contained 55, of the 272 polypeptides visualized in these fractions. Two-dimensional electrophoresis combined with subcellular fractionation may be useful tools for examining changes in polypeptide composition caused by nitrogen starvation.     

Indicators of Postoperative Pain in Syrian Hamsters (Mesocricetus auratus)

Despite the use of Syrian hamsters (Mesocricetus auratus) in research, little is known about the evaluation of pain in this species. This study investigated whether the frequency of certain behaviors, a grimace scale, the treat-take-test proxy indicator, body weight, water consumption, and coat appearance could be monitored as signs of postoperative pain in hamsters in a research setting. Animals underwent no manipulation, anesthesia only or laparotomy under anesthesia. An ethogram was constructed and used to determine the frequencies of pain, active and passive behaviors by in-person and remote videorecording observation methods. The Syrian Hamster Grimace Scale (SHGS) was developed for evaluation of facial expressions before and after the surgery. The treat-take-test assessed whether surgery would affect the animals’ motivation to take a high-value food item from a handler. The hypothesis was that behavior frequency, grimace scale, treat-take-test score, body weight, water consumption, and coat appearance would change from baseline in the surgery group but not in the no-intervention and anesthesia-only groups. At several time points, pain and passive behaviors were higher than during baseline in the surgery group but not the anesthesia-only and no-intervention groups. The SHGS score increased from baseline scores in 3 of the 9 animals studied after surgery. The frequency of pain behaviors and SHGS scores were highly specific but poorly sensitive tools to identify animals with pain. Behaviors in the pain category were exhibited by chiefly, but not solely, animals that underwent the laparotomy. Also, many animals that underwent laparotomy did not show behaviors in the pain category. Treat-take-test scores, body weight, water consumption, and coat appearance did not change from baseline in any of the 3 groups. Overall, the methods we tested for identifying Syrian hamsters experiencing postoperative pain were not effective. More research is needed regarding clinically relevant strategies to assess pain in Syrian hamsters.

Pain experienced by laboratory animals can affect both animal welfare and research results. Little is known about the evaluation of pain in Syrian hamsters (Mesocricetus auratus) in the laboratory setting. However, various research models using Syrian hamsters involve surgery and are presumed to cause pain.^16,47,49 In 2018 alone, the USDA reported that 35,695 hamsters were used for research studies involving painful procedures.⁴⁸ Previously published behaviors exhibited by hamsters in response to pain include hunched posture with head down, reluctance to move, increased depression or aggression, extended sleep periods, and weight loss.^7,8,10,16,21 How these behaviors are affected by factors such as the type of painful stimulus, anesthetic protocol, handling procedures, and environmental conditions is unclear. The practicality of observing these signs in the research environment is uncertain and likely complicated by the nocturnal nature of Syrian hamsters and an assumed propensity of this species to mask pain, much like other prey species.^8,14,16A significant need exists for published data investigating whether behavioral observations or other clinical indicators can help recognize, quantify, or monitor pain in hamsters in a research setting. Detailed behavioral observations and well-controlled studies are needed to develop a system to assess postoperative pain in laboratory animals.^8,33 Moreover, little information is available on the efficacy of analgesic agents in hamsters.¹ The few studies of analgesics in hamsters rely on the mitigation of evoked pain responses (such as using a hot plate), which has limited relevance to clinical situations such as postoperative pain.^8,32,36,51 To date, no published literature has evaluated the efficacy or safety of analgesics to treat postoperative pain in hamsters. Validated real-time and practical methods for evaluating pain in Syrian hamsters would support the evaluation of analgesic efficacy in this species.Various assessments have been developed to identify signs of pain in other species. Behavioral ethograms have been used to evaluate pain and analgesic efficacy in mice, rats, rabbits, and guinea pigs in the research environment.^{5,6,20,23,25,34,35,39-41,53} Another tool used to evaluate pain in animals is the grimace scale, which has been developed for mice, rats, rabbits, ferrets, cats, sheep, pigs, horses, and even harbor seals.^{3,4,9,11,13,15,19,22,26,30,37,45,50} The use of a proxy indicator, such as burrowing and time-to-integrate-to-nest in mice and time-to-consume in guinea pigs, can be used as an additional tool for the evaluation of pain.^{5,17,18,35,38}Because none of the previously mentioned assessment techniques were specific to hamsters, we here explored using these approaches to detect pain in Syrian hamsters that underwent laparotomy in a laboratory setting. We developed a species-specific ethogram and the Syrian Hamster Grimace Scale (SHGS). We also devised a novel proxy indicator of pain for use in Syrian hamsters, the treat-take-test (TTT), which is based on hamsters’ natural behavior to hoard food.^16,46,49,52 Although water intake, body weight, and coat appearance are non-specific indicators of pain, we also measured these parameters.^5,19,23,33 Furthermore, we analyzed the effects of the presence of an observer and time of day. We hypothesized that behavior frequency, grimace scale, treat-take-test score, body weight, water consumption, and coat appearance would change from baseline in the surgery group but not in the no-intervention and anesthesia-only groups.