Angiographic, Anatomic and Clinical Technique Descriptions of A Subcarapacial Venipuncture Site for Chelonians

Transcription

1 C l i n i c a l T e c h n i q u e s Angiographic, Anatomic and Clinical Technique Descriptions of A Subcarapacial Venipuncture Site for Chelonians Sonia M. H ernandez-d ivers1, DVM, Stephen J. Hernandez-D ivers2, BVetMed, D ZooM ed (Reptilian), M RCVS, RCVS Specialist in Zoo & W ildlife M edicine (Reptiles), Jeanette W yneken3, PhD 1. Division of Wildlife Health, Department of Clinical Sciences, Division of Wildlife Health, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, USA Author s current address: Exotic Animal Wildlife and Zoo Animal Medicine, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, University of Georgia, Athens, GA , USA 2. Exotic Animal, Wildlife and Zoo Animal Medicine, Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA 3. Department of Biological Sciences, Florida Atlantic University, 777 Glades Road, Boca Raton, FL 33431, USA A bstr a c t: A novel technique for chelonian venipunture is described. Angiographic studies using radioopaque iohexol, static radiography, and video fluoroscopy were undertaken in several chelonian species. Latex injected dissections were also performed to confirm the vascular anatomy. These studies indicate that a clinically useful venipuncture site is present in the anterior dorsal midline at the junction of the common intercostal veins and the caudal cervical branch of the external jugular veins, located just craniad of the last mobile cervical vertebra. K ey W o r d s: Chelonia, vein, venipuncture, blood collection, anatomy, radiography, fluoroscopy. INTRODUCTION The ability to safely and effectively collect blood from reptiles is a vital aspect of herpetological medicine. Blood may be collected from tortoises, turtles and terrapins from the jugular veins, brachial veins, femoral veins, ventral, dorsal or lateral coccygeal veins, post-occipital venous sinus, heart, and trimmed toenails (Owens and Ruiz, 1980, Frye, 1991, Jacobson, et al, 1992, Jacobson, 1993, Jenkins, 1996, Kolle and Hoffman, 1996, Murray, 2000). The preferred method of blood collection often depends upon the species (size, disposition), the volume required and the preference of the clinician. Less humane methods of collection, such as clipping toenails, are no longer ethically acceptable. Many blood collection sites pose difficulties (Murray, 2000). For example, venipuncture of the coccygeal vessels often results in lymphatic contamination, while jugular venipuncture requires extension of the head and neck, which can be difficult in animals that have not been chemically restrained. In addition, blood collection from peripheral vessels may yield small volumes. Practical experience has indicated that a subcarapacial venipuncture site has clinical value in chelonians. Depending upon the species and conformation of the carapace, the needle may be inserted anteriorly along the midsaggital line at an angle up to 60 just caudal to the skin-carapace junction. The needle is advanced in a caudodorsal direction towards the cranial aspect of the eighth cervical vertebra, maintaining slight negative pressure (Figure 1). If a vertebra is encountered, the needle is withdrawn slightly and redirected further craniad or caudad. This approach has been used successfully by two of the authors (SMHD and SJHD) for the collection of blood from North African tortoises, Testudo graeca, Herman s tortoises, Testudo hermanni, Russian tortoises, Testudo horsfieldi, African spurred tortoise, Geochelone sulcata, leopard tortoises, Geochelone pardalis, red-footed tortoises, Geochelone carbonaria, South American yellow-footed tortoises, Geochelone denticulate, red-eared sliders, Trachemys scripta elegans, snapping turtles, Chelydra serpentina, sliders, Trachemys picta, eastern box turtles, Terrapene c. Carolina, ornate box turtles, Terrapene ornata, spotted turtles, Clemmys guttata, Blanding s turtles, Emydoidea blandingi, radiated tortoises, Geochelone radiata, and African hinged-back tortoises, Kinixys belliana. The particular advantages of the subcarapacial site include large volume collection, and accessibility in very small and large specimens whether the chelonian head is extended or retracted (Figures 2, 3, 4, 5). In an attempt to accurately describe this venous site, a detailed review of the chelonian anatomy literature was undertaken (Bojanus, 1819, Noble and Noble, 1909, Thompson, 1932, Ashley, 1975, Owens and Ruiz, 1980). In chelonians, the external jugular veins course caudally along the dorsolateral 32 Journal of Herpetological Medicine and Surgery Volume 12, No. 2,2002

2 Anastomosis of the common intercostal veins Needle External jugular vein Caudal cervical branch of jugular veins Figure 1. Diagram illustrating the relationship between the carapace, cervical spine and venous circulation of a chelonian, including the placement of the needle for venipuncture (Adapted ffombojanus, 1819). aspect of either side of the neck. There are several cervical branches that connect the two external jugular veins transversely across the dorsum of the neck, deep to the major dorsal muscles (Figure 6). The most caudal cervical branch joins the cranial anastomosis of the common intercostal vessels that arise from the azygous veins (Figure 7). The caudal aspect of each external jugular vein receives branches ventromedially from the esophagus and the internal jugular, and laterally from the subclavian and brachial veins of the pectoral region. The junction of the common intercostals and the caudal branch from the external jugulars is along the dorsal midline, just craniad of the eighth cervical vertebra. This was confirmed as the site of venipuncture using angiography and latex injection of the venous circulation in cadavers. This paper describes the anatomical location of the subcarapacial venipuncture site through radiographic, fluoroscopic imaging and cadaver dissection to facilitate its clinical use in chelonians. MATERIALS AND METHODS The subcarapacial vasculature was assessed by static radiographic angiography, fluoroscopy, and dissection of latex-injected cadavers. Radiography and Fluoroscopy - Case 1- An adult captive, 776 g, female red-eared slider, Trachemys scripta elegans, of unknown age, considered clinically normal and in good body condition, was used for static radiographic angiography. Prior to and after the radiographic study, the slider was maintained at 27 C (80 F). During the procedure, the animal was maintained on a heating pad (Orthobionics, Inc. Dallas, TX). A doppler unit (Ultrasonic Doppler Flow Detector 811-B, Parks Medical Electronics Inc., Aloha, OR) was used to measure heart rate, and cloacal temperature was recorded before the procedure. W hole body survey radiographs, including dorsoventral, horizontal lateral and horizontal craniocaudal views, were taken prior to the administration of the contrast agent. The slider was manually restrained in a normal horizontal orientation with the two forelimbs secured by a handler. The head was gently forced into the shell using an empty 3 ml syringe case. A 3 ml syringe with a 2.5 cm (1 in) 2 2 ga needle was positioned caudal to the cranial rim of the carapace and carefully advanced caudodorsally, approximately 45 above the the horizontal plane, towards the eighth cervical vertebra. Upon the aspiration of blood, the syringe was disconnected and the blood submitted for hematocrit evaluation. A second syringe containing 1.5 ml iohexol (407 mg I/kg; Omnipaque, 350 mg I/ml, Nicomed Inc, 101 Carnegie Center, Princeton, NJ) was attached to the needle, and the contrast injected over three seconds. The injection was immediately followed by dorsoventral and lateral radiographs, and a second series of radiographs were taken 15 min later. No adverse effects were noted until observations ceased, 30 d after the procedure. Case 2 - A free-ranging, 560 g, male red-eared slider, Trachemys scripta elegans, was presented for treatment of minor shell injuries. Three weeks after these injuries were repaired, the animal was deemed in good health. This turtle was handled in the same manner as Case 1. The animal was similarly restrained, with the head gently forced into the shell. A 3 ml syringe with a 2.5 cm (1 in) 22 ga needle was filled with 0.75 ml of iohexol (147 mg I/kg). The needle was advanced on the dorsal midline, just caudal to the junction of the skin and the ventral carapace, approximately 30 to the horizontal towards the eighth cervical vertebra. The needle was advanced with negative pressure until bone was felt, then withdrawn slightly until a flash of blood contaminated with clear fluid was present in the needle hub. At that time, the iohexol was injected into the site over five seconds and the needle removed. The turtle was positioned under a fluoroscopy unit and dorsoventral and lateral radiographic images were recorded. The procedure was also recorded onto videotape. The time of injection, as well as the time of each radiograph was recorded. No ill effects were observed and the animal was released to the care of a rehabilitator one week later. Case 3 - An adult, 485 g, female eastern box turtle, Terrapene c. Carolina, of unknown age underwent the same fluoroscopic procedure as described previously. The animal was restrained and the heart rate and body temperature were measured as described in the previous cases. The syringe was advanced as previously described; however, in this case, 0.1 ml of blood was aspirated prior to injection to ensure intravascular administration. This turtle received 0.7 ml of iohexol (118.8 mg I/kg) over five seconds. The animal was placed under the fluoroscopy unit and several dorsoventral and lateral radiographic images were recorded. The procedure was also recorded onto videotape. No untoward effects were observed and the animal was released to its owners. Cadaver Dissection and Latex Injection - Dissection and latex injection was performed on six chelonians. Two adult female sliders, Trachemys scripta elegans, and one juvenile gopher tortoise, Gopherus polyphemus, were obtained following euthanasia. Two dead green turtles, Chelonia mydas, and one dead hawksbill, Eretmochelys imbricata, were obtained from the Florida Department of Marine Research Sea Turtle Volume 12, No. 2,2002 Journal of Herpetological Medicine and Surgery 33

3 34 Figure 2. Blood collection from the subcarapacial site of an adult red-eared terrapin, Trachemys scripta elegans, with the head retracted. Figure 4. Blood collection from the subcarapacial site of an adult box turtle, Terrapene c. Carolina, with the head retracted. A plastic gag has been used to prevent the complete closure of the cranial plastron and permit the introduction of the needle. Figure 3. Blood collection from the subcarapacial site of a juve nile Greek tortoise, Testudo graeca, with the head extended. Figure 5. Blood collection from the subcarapacial site of an African spurred tortoise, Geochelone sulcata, with the head retracted. Stranding and Salvage Network (JW permit #073). The dorsal and lateral skin of these turtles was removed from the neck and from the anterior shoulders from the carapace toward the plastron. The external jugular veins were exposed by blunt dissection of the overlying connective tissue, fat, and con strictor colli muscles. A 1 ml syringe was filled with blue latex injection medium (Carolina Biological Supply Co., 2700 York Road, Burlington, NC) and the end fitted with 2.5 cm (1 ) or 1.25 cm (0.5 ) 27 ga needles as required. The external jugular veins were then injected with the latex along their length. The latex was cured by injection of small amounts of acetic acid. RESULTS Journal of Herpetological Medicine and Surgery Case 1 - Doppler auscultation was unremarkable. Heart rate of 48 bpm was recorded at a cloacal temperature of 26 C (80 F). Blood collected from the subcarapacial site yielded a packed cell volume of 28%. Radiography immediately after contrast administration demonstrated a dilated sinus just cra nial to the eighth cervical vertebra. Contrast material was noted to pass ventrally in the caudal cervical branch and caudoventrally in the external jugular veins and vena cava. Contrast material was also detected within the heart. Further imaging of the area 15 min later failed to indicate any con trast in the region. Case 2 - The heart rate of this turtle was 44 bpm. The body temperature was 31 C (8 8 F). The contrast material injected into the subcarapacial site remained in a well-circumscribed area just cranial to the eighth cervical vertebra (Figure 8 ) for Volume 12, No. 2,2002

4 Figure 6. Lateral view of the venous circulation of the cranial carapace of a chelonian depicting the relationship between the caudal transverse cervical branches of the external jugular veins and the cranial anastomosis of the common intercostal vessels that arise from the azygous veins (Adapted from Bojanus, 1819). Cranial over 45 min. The video recording failed to demonstrate any significant movement of the material, even after two hours post-injection. Case 3 - The heart rate of this turtle was 32 bpm and the body temperature was 27 C (80 F). The first lateral radi ograph showed the presence of the contrast material at the level of the eighth cervical vertebra, coursing ventrally to the external jugular vein and then caudoventrally in the vena cava towards the heart (Figure 9). The contrast material filling the ventricle was visualized on video as the ventricle contracted at approximately 30 beats per minute. The corresponding dorsoventral view showed the contrast material at the level of the eighth cervical vertebra and coursing caudally (Figure 10). Ten minutes post-injection, contrast material was no longer visible on either view. Cadaver Dissection and Latex Injection - Latex injection showed the well-developed external jugular veins, the caudal cervical branch which connect the external jugular veins, and the common intercostal veins, in all species dissected. The anastomoses of the common intercostals and the caudal cervi cal branch of the external jugular veins were more developed in the tortoise dissected, but less prominent in the emydid tur tles, Trachemys spp., and sea turtles, Chelonia spp., and Eretmochelys spp. The connection between the common inter costals and caudal cervical branch of the external jugulars along the dorsal midline was obvious and clearly visible (Figure 11). DISCUSSION Caudal cervical branch of jugular veins Common intercostal veins S i Azygous veins Figure 7. Dorsal view of the venous circulation of the cranioventral dorsal carapace of a chelonian depicting the relationship between the caudal transverse cervical branch of the external jugular veins and the cranial anastomosis of the com mon intercostal vessels that arise from the azygous veins (Adapted from Bojanus, 1819). Volume 12, No. 2,2002 Case 2 - The contrast material in this case was injected into a well-circumscribed region just cranial to the eighth cervical vertebrae. Judging by contamination of the blood aspirated with a clear fluid prior to injection, it is presumed that a lym phatic sinus is in close association to the venipuncture site. The contrast material remained in this region for over 45 min, further supporting that this material was extravasated. The experience of two authors (SMHD and SJHD) when sampling other individuals further suggests that lymphatic contamina tion is possible if the needle is not advanced sufficiently caudodorsally, to the level of the eighth cervical vertebrae. Cases 1 and 3 - The contrast material was seen entering a small region at the level of the eighth cervical vertebra imme diately post-injection. The material coursed caudoventrally and reached the heart within three to five seconds of adminis tration. Hence, the material is presumed to have entered the communication of the junction of the common intercostal ves sels and the caudal transverse cervical branch of the external jugular veins. The aspiration of whole blood further supports administration into the vascular compartment. The contrast agent then coursed through the jugular veins, the vena cava and into the heart. Anatomical dissections indicate that this novel venipunc ture site is formed by the junction of the common intercostals and the caudal cervical branch of the external jugular veins. This site can be a useful alternative method of venipuncture for the practitioner, if the technique is followed carefully. With little practice, large volumes of whole blood can be col lected from small or uncooperative individuals without the need for chemical restraint. The authors (SMHD, SJHD) have also used this site for the successful administration of intra venous agents. The primary problem with this technique is Journal of Herpetological Medicine and Surgery 35

5 Figure 8. Dorsoventral radiographic view of the red-eared slider, Trachemys scripta elegans, described in case 2. Radio-opaque contrast material (arrows) is seen pooling cranial to the eighth cervical vertebra. Cranial Figure 10. Dorsoventral radiographic view of the eastern box turtle, Terrapene c. Carolina, described in case 3, within ten seconds of iohexol administration. The site of injection is depicted by X. The arrows outline the radio-opaque contrast material coursing caudoventrally in the external jugular veins and vena cava towards the heart. Figure 9. Horizontal beam lateral radiographic view of the eastern box turtle, Terrapene c. Carolina, described in case 3, within ten seconds of iohexol administration. The radio opaque contrast material is seen coursing from the site of injection ( 1), to the caudal cervical branch of the external jugulars (2), to the external jugulars (3), and into the heart (4). Figure 11. Latex injected dissection of a gopher tortoise, Gopherus polyphemus, with the dorsal skin of the neck and the cranial carapace removed. Note the right external jugular vein ( 1), caudal cervical branch of the external jugular veins (2), venipuncture site at the junction of the common intercostals and caudal cervical branch (3), left common intercostal vein (4), right common intercostal vein (5), right azygous vein (6 ), and the eighth cervical vertebra (7). 36 Journal of Herpetological Medicine and Surgery Volume 12, No. 2,2002

6 inadvertent aspiration of lymph cranial to the recommended site, as evidenced by case two. This is of particular importance in large animals where commonly used 2.5 cm (1 in) 22 ga needles may be of insufficient length to reach the described venipuncture site. In such cases, 3.8 cm (1.5 in) hypodermic needles or 5 cm (2 in) spinal needles may be necessary. If lymph is aspirated, it could affect results of a biochemical profile. Other than the measurement of the hematocrit in Case 1, the hematological or biochemical parameters of blood collected from this site were not investigated in this study. Although clinically the authors have used this site for sample collection for complete blood cell counts and biochemical analyses, further investigations to correlate samples gathered from this site with those from a jugular or dorsal venous sinus samples are needed. The curvature of the carapace is also a significant factor in reaching this site, and chelonians with highly domed carapaces may also require longer needle lengths for successful sample collection. Clinicians are advised to pre-measure the length of the needle against the carapace, to mentally visualize the regional skeletal and vascular anatomy, and aim for the eighth cervical vertebra. ACKNOWLEDGEMENTS The authors would like to thank the Radiology Department at Cornell University s College of Veterinary Medicine, and in specific, Dr. Tina Wilson, for their contribution with the fluoroscopy studies. We also thank Drs. George V. Kollias for his guidance with this technique and Ale Aguirre for his assistance with the fluoroscopy studies. R. Funk, P. Lutz, K. Singel, and W. Teas supplied carcasses for dissection. Sea Turtle dissections were conducted under Florida Permit #073. REFERENCES Ashley LM Laboratory Anatomy of the Turtle. WC Brown, Dubuque. Bojanus LH Anatomie Testudinis Europaeae. Tipographi Universitatis, Vilnae, Lithuania. Frye, FL Hematology as applied to clinical reptile medicine. In (Frye FL (ed): Biomedical and Surgical Aspects of Captive Reptile Husbandry. Krieger, Malabar, FL: Jacobson ER Blood collection techniques in reptiles: Laboratory investigations. In Fowler ME (ed): Zoo and Wild Animal Medicine: Current Therapy 3. WB Saunders, Philadelphia,PA: Jacobson ER, Schumacher J, Green M Field and clinical techniques for sampling and handling blood for hematologic and selected biochemical determinations in the desert tortoise (Xerobates agassizii). Copeia, 1:237. Jenkins JR Diagnostic and clinical techniques. In Mader DR (ed): Reptile Medicine and Surgery. WB Saunders, Philadelphia, PA: Kolle P, Hoffman R Blutparameter als Hilfe in der Diagnostik von Reptilienkrankheiten. Tierarztl Prax 24: Murray MJ Reptilian blood sampling and artifact considerations. In Fudge AM (ed): Laboratory Medicine Avian and Exotic Pets. WB Saunders, Philadelphia, PA: Noble GA, Noble ER A brief anatomy of the turtle. Stanford University, California. Owens DW, Ruiz GJ New methods of obtaining blood and cerebrospinal fluid from marine turtles. Herpetologica, 36:17. Thompson JS The anatomy of the tortoise. The Scientific Proceedings of the Royal Dublin Society. 20 (28): Volume 12, No. 2,2002 Journal of Herpetological Medicine and Surgery 37