Harnessing cerebrospinal fluid circulation for drug delivery to brain tissues.

Initially thought to be useful only to reach tissues in the immediate vicinity of the CSF circulatory system, CSF circulation is now increasingly viewed as a viable pathway to deliver certain therapeutics deeper into brain tissues. There is emerging evidence that this goal is achievable in the case of large therapeutic proteins, provided conditions are met that are described herein. We show how fluid dynamic modeling helps predict infusion rate and duration to overcome high CSF turnover. We posit that despite model limitations and controversies, fluid dynamic models, pharmacokinetic models, preclinical testing, and a qualitative understanding of the glymphatic system circulation can be used to estimate drug penetration in brain tissues. Lastly, in addition to highlighting landmark scientific and medical literature, we provide practical advice on formulation development, device selection, and pharmacokinetic modeling. Our review of clinical studies suggests a growing interest for intra-CSF delivery, particularly for targeted proteins.

Automated Blood Sampling in a Canine Telemetry Cardiovascular Model.

Successful implementation of automated blood sampling (ABS) into a telemetry instrumented canine cardiovascular model provides simultaneous cardiovascular assessment of novel compounds while collecting multiple blood samples for analysis of drug level, cytokines, and biomarkers. Purpose-bred male Beagle dogs (n = 36) were instrumented with a dual-pressure telemetry transmitter and vascular access port. Modifications to acclimation practices, surgical procedures, and housing were required for implementation of ABS in our established cardiovascular canine telemetry colony. These modifications have increased the use and reproducibility of the model by combining early pharmacokinetic and cardiovascular studies, thus achieving both refinement and reduction from a 3R perspective. In addition, the modified model can shorten timelines and reduce the compound requirement in early stages of drug development. This telemetry-ABS model provides an efficient means to quickly identify potential effects on key cardiovascular parameters in a large animal species and to obtain a more complete pharmacokinetic-pharmacodynamic profile for discovery compounds.

The cardiopulmonary effects of anesthetic induction with isoflurane, ketamine-diazepam or propofol-diazepam in the hypovolemic dog.

OBJECTIVE: To evaluate and compare the cardiopulmonary effects of induction of anesthesia with isoflurane (Iso), ketamine-diazepam (KD), or propofol-diazepam (PD) in hypovolemic dogs. Study design Prospective randomized cross-over trial. ANIMALS: Six healthy intact, mixed breed, female dogs weighing 20.7 +/- 4.2 kg and aged 22 +/- 2 months. Methods Dogs had 30 mL kg(-1) of blood removed at a rate of 1.5 mL kg(-1) minute(-1) under isoflurane anesthesia. Following a 30-minute recovery period, anesthesia was reinduced. Dogs were assigned to one of three treatments: isoflurane via facemask using 0.5% incremental increases in the delivered concentration every 30 seconds, 1.25 mg kg(-1) ketamine and 0.0625 mg kg(-1) diazepam intravenously (IV) with doses repeated every 30 seconds as required, and 2 mg kg(-1) propofol and 0.2 mg kg(-1) diazepam IV followed by 1 mg kg(-1) propofol increments IV every 30 seconds as required. Following endotracheal intubation all dogs received 1.7% end-tidal isoflurane in oxygen. Cardiopulmonary variables were recorded at baseline (before induction) and at 5 or 10 minute intervals following endotracheal intubation. RESULTS: Induction time was longer in Iso (4.98 +/- 0.47 minutes) compared to KD (3.10 +/- 0.47 minutes) or PD (3.22 +/- 0.45 minutes). To produce anesthesia, KD received 4.9 +/- 2.3 mg kg(-1) ketamine and 0.24 +/- 0.1 mg kg(-1) diazepam, while PD received 2.2 +/- 0.4 mg kg(-1) propofol and 0.2 mg kg(-1) diazepam. End-tidal isoflurane concentration immediately following intubation was 1.7 +/- 0.4% in Iso. Arterial blood pressure and heart rate were significantly higher in KD and PD compared to Iso and in KD compared to PD. Arterial carbon dioxide partial pressure was significantly higher in PD compared to KD and Iso immediately after induction. CONCLUSIONS AND CLINICAL RELEVANCE: In hypovolemic dogs, KD or PD, as used in this study to induce anesthesia, resulted in less hemodynamic depression compared to isoflurane.

Systemic innate immune responses following intrapulmonary delivery of CpG oligodeoxynucleotides in sheep.

Mucosal delivery of CpG oligodeoxynucleotide (ODN) in mice has been shown to induce potent innate immunostimulatory responses and protection against infection. We evaluated the efficacy of CpG ODN in stimulating systemic innate immune responses in sheep following delivery to the pulmonary mucosa. Intrapulmonary (IPM) administration of B-Class CpG ODN in saline induced transient systemic responses which included increased rectal temperatures, elevated serum 2'5'-A synthetase and haptoglobin concentrations. The ODN dose required to induce detectable systemic responses following IPM delivery could be reduced by approximately 80% if the CpG ODN was administered in 30% emulsigen instead of saline. Intrapulmonary B-Class CpG ODN formulated in 30% emulsigen produced similar effects when compared to those seen following SC injection. These responses were CpG ODN-specific since control GpC ODN did not induce any detectable response. Intrapulmonary administration of both B-Class and the newly described C-Class CpG ODN produced similar effects indicating that both classes of CpG ODN were comparably effective in stimulating innate immune system following mucosal delivery. Administration of CpG ODN directly into the lungs or delivery of CpG ODN via an intratracheal (IT) infusion also produced similar systemic responses. These observations support the conclusion that mucosal delivery of CpG ODN is an effective route for induction of systemic acute phase responses and antiviral effector molecules in large animals, and may be helpful in controlling systemic infections.

Cardiopulmonary effects of administration of a combination solution of xylazine, guaifenesin, and ketamine or inhaled isoflurane in mechanically ventilated calves.

OBJECTIVE: To compare the cardiopulmonary effects of administration of a solution of xylazine, guaifenesin, and ketamine (XGK) or inhaled isoflurane in mechanically ventilated calves undergoing surgery. ANIMALS: 13 male calves 2 to 26 days of age. Procedures-In calves in the XGK group, anesthesia was induced (0.5 mL/kg) and maintained (2.5 mL/kg/h) with a combination solution of xylazine (0.1 mg/mL), guaifenesin (50 mg/mL), and ketamine (1.0 mg/mL). For calves in the isoflurane group, anesthesia was induced and maintained with isoflurane in oxygen. The rates of XGK infusion and isoflurane administration were adjusted to achieve suitable anesthetic depth. All calves received 100% oxygen and were mechanically ventilated to maintain end-tidal carbon dioxide concentrations from 35 to 40 mm Hg and underwent laparoscopic bladder surgery through an abdominal approach. Cardiopulmonary variables were measured before induction and at intervals up to 90 minutes after anesthetic induction. RESULTS: The quality of induction was excellent in all calves. The XGK requirements were 0.57 +/- 0.18 mL/kg and 2.70 +/- 0.40 mL/kg/h to induce and maintain anesthesia, respectively. Heart rate was significantly lower than baseline throughout the anesthetic period in the XGK group. Systolic arterial blood pressure was significantly higher in the XGK group, compared with the isoflurane group, from 5 to 90 minutes. Cardiac index was lower than baseline in both groups. Differences between groups in cardiac index and arterial blood gas values were not significant. CONCLUSIONS AND CLINICAL RELEVANCE: Administration of XGK resulted in excellent anesthetic induction and maintenance with cardiopulmonary alterations similar to those associated with isoflurane in mechanically ventilated calves.

Comparison of hemodynamic, clinicopathologic, and gastrointestinal motility effects and recovery characteristics of anesthesia with isoflurane and halothane in horses undergoing arthroscopic surgery.

OBJECTIVE: To compare hemodynamic, clinicopathologic, and gastrointestinal motility effects and recovery characteristics of halothane and isoflurane in horses undergoing arthroscopic surgery. ANIMALS: 8 healthy adult horses. PROCEDURE: Anesthesia was maintained with isoflurane or halothane (crossover study). At 6 intervals during anesthesia and surgery, cardiopulmonary variables and related derived values were recorded. Recovery from anesthesia was assessed; gastrointestinal tract motility was subjectively monitored for 72 hours after anesthesia. Horses were administered chromium, and fecal chromium concentration was used to assess intestinal transit time. Venous blood samples were collected for clinicopathologic analyses before and 2, 24, and 48 hours after anesthesia. RESULTS: Compared with halothane-anesthetized horses, cardiac index, oxygen delivery, and heart rate were higher and systemic vascular resistance was lower in isoflurane-anesthetized horses. Mean arterial blood pressure and the dobutamine dose required to maintain blood pressure were similar for both treatments. Duration and quality of recovery from anesthesia did not differ between treatments, although the recovery periods were somewhat shorter with isoflurane. After isoflurane anesthesia, gastrointestinal motility normalized earlier and intestinal transit time of chromium was shorter than that detected after halothane anesthesia. Compared with isoflurane, halothane was associated with increases in serum aspartate transaminase and glutamate dehydrogenase activities, but there were no other important differences in clinicopathologic variables between treatments. CONCLUSIONS AND CLINICAL RELEVANCE: Compared with halothane, isoflurane appears to be associated with better hemodynamic stability during anesthesia, less hepatic and muscle damage, and more rapid return of normal intestinal motility after anesthesia in horses undergoing arthroscopic procedures.

Subcutaneous, but not intratracheal administration of the TLR9 agonist, CpG DNA transiently reduces parainfluenza-3 virus shedding in newborn lambs.

Synthetic oligodeoxynucleotides (ODN) containing CpG motifs signal through TLR9 and activate innate immunity resulting in protection against a variety of parasitic, bacterial and viral pathogens in mouse models. However, few studies have demonstrated protection in humans and large animals. In the present investigations, we evaluated protection by CpG ODN in a parainfluenza-3 (PI-3) virus infection in neonatal lambs. Subcutaneous (SC) injection of CpG ODN induced high levels of 2'5'-A synthetase and significantly reduced PI-3 virus shedding in newborn lambs. Furthermore, pre-treatment of newborn lambs with SC CpG ODN 2 days, but not 6 days prior to the virus challenge was protective. In contrast, intratracheal (IT) administration of CpG ODN induced 2'5'-A synthetase but had no significant impact on PI-3 virus shedding in nasal secretions. We conclude that a systemic administration of CpG ODN and the timing of the treatment are critical for the protection of neonatal lambs against a respiratory viral infection.

Oral DNA immunization in the second trimester fetal lamb and secondary immune responses in the neonate.

We previously demonstrated that oral DNA vaccination of the third trimester fetus may be an effective strategy to prevent vertical disease transmission despite an immature immune system and the possibility of developing tolerance. The present investigation examined oral DNA vaccine delivery and immunogenicity in the second trimester fetal lamb (50-100 days of gestation (dg)). Histological examination revealed nucleated cells in the superficial layers of the oral mucosa. Therefore, luciferase-encoding DNA plasmid was injected into the oral cavity of 65-70 dg fetuses to monitor plasmid expression. Luciferase activity was detected in the oral mucosa of all fetuses but the level of luciferase activity varied among individual fetuses. Luciferase activity was also detected within the tonsils and lymph nodes draining the oral cavity. Oral DNA immunization between 55 and 83 dg with a truncated glycoprotein D (tgD)-encoding plasmid induced germinal centres in the draining lymph nodes and detectable tgD-specific serum antibody titers and/or IFNgamma-secreting cell responses in 16 of 24 (67%) fetuses. The tgD-specific antibody and IFNgamma responses persisted until birth in some fetuses but the magnitude of antibody titers in second trimester fetuses was low relative to antibody titers induced following oral DNA immunization in the third trimester. Lambs immunized during the second trimester of gestation responded to neonatal DNA immunization and anamnestic responses were detected in some lambs immunized as early as 67-72 dg. These observations confirmed that oral DNA immunization of the early second trimester fetus induced antigen-specific immune responses with no evidence of tolerance induction.

The effect of hypovolemia due to hemorrhage on the minimum alveolar concentration of isoflurane in the dog.

OBJECTIVE: To determine the effect of hypovolemia on the minimum alveolar concentration (MAC) of isoflurane in the dog. STUDY DESIGN: Randomized, cross-over trial. ANIMAL POPULATION: Six healthy intact mixed breed female dogs weighing 18.2-29.0 kg. METHODS: Dogs were randomly assigned to determine the MAC of isoflurane in a normovolemic or hypovolemic state with a minimum of 18 days between trials. On both occasions, anesthesia was initially induced and maintained for 40 minutes with isoflurane delivered in oxygen while vascular catheters were placed in the cephalic vein and dorsal metatarsal artery. In dogs assigned to the hypovolemic group, 30 mL kg(-1) of blood was removed at 1 mL kg(-1) minute(-1) from the arterial catheter. All dogs were allowed to recover from anesthesia. Thirty minutes after the discontinuation of isoflurane, anesthesia was re-induced with isoflurane in oxygen delivered by face mask. The tracheas were intubated, and connected to an anesthetic machine with a Bain anesthetic circuit. Mechanical ventilation was instituted at a rate of 10 breaths minute(-1) with the tidal volume set to deliver 10-15 mL kg(-1). Airway gases were monitored continuously and tidal volume was adjusted to maintain an end-tidal carbon dioxide level of 35-40 mmHg (4.67-5.33 kPa). Body temperature was maintained at 37-38 degrees C (98.6-100.4 degrees F). The MAC determination was performed using an electrical stimulus applied to the toe web and MAC was defined as the mean value of end-tidal isoflurane between the concentrations at which a purposeful movement did and did not occur in response to the electrical stimulus. The MAC values were compared between groups using a Student's t-test. RESULTS: The MAC of isoflurane was significantly less in hypovolemic dogs (0.97 +/- 0.03%) compared with normovolemic dogs (1.15 +/- 0.02%) (p < 0.0079). CONCLUSIONS AND CLINICAL RELEVANCE: The MAC of isoflurane is reduced in dogs with hypovolemia resulting from hemorrhage. Veterinarians should be prepared to deliver a lower percentage of isoflurane to maintain anesthesia in hypovolemic dogs during diagnostic and therapeutic procedures.