Implantation Of Guide Cannula Research Articles

Biocompatibility and customizability: Expanding possibilities with 3D printed guide cannulas

BackgroundIntracerebral cannulation bypasses the blood-brain barrier, and is frequently used for targeted drug delivery to specific brain structures. Despite the availability of brain infusion kits and manual injections without cannulation, the traditional design of guide cannulas continues to be utilized in research. Several protocols describing guide cannula manufacture from stainless steel needles have been published previously. New methodWe describe a method for producing the first fully plastic guide cannula intended for intracerebroventricular injections in mice using Dental Sand A1-A2 resin and digital light processing 3D printing. ResultsThe lack of resin neurotoxicity for primary rat cortical neuron cultures was shown. Histological evaluations performed 6 weeks after guide cannula implantation to C57/black mice show that plastic cannula are biocompatible. Microglial and astroglial reactions to plastic cannulas are reduced compared to lab-made stainless steel cannulas. Plastic cannulas are less prone to obstruction, and remained unobstructed over the course of 3 weeks of daily injections, while 50 % of stainless steel cannula became impassable by the 2 week mark. Comparison with existing methodsThese are the first published cannulas intended for applications in mice which combine the presence of usable threads, allowing dummy cannula fixation, with a low profile and small footprint compared to commercially available cannulas. ConclusionsEditable parametric and stl files for reproducing the cannulas presented in this manuscript are included. The method described in this paper is accessible to most laboratories, enabling near-perfect standardization in length combined with a high level of customizability.

Serotonin as a Regulator of Leptin-Mediated Food Intake Control Within a Novel Neuronal Circuit Between the Hypothalamus and Raphe Nuclei

The onset and exacerbation of obesity involves the overproduction of the adipocyte-derived hormone leptin, a key mediator of homeostatic appetite regulation and a signal for satiety. Although leptin’s hypothalamic regulation of food intake has been extensively investigated, its role in tandem with the anorectic neurotransmitter serotonin (5-HT) has been less characterized. 5-HT is synthesized in the dorsal raphe nucleus (DRN) where anatomical projections to many hypothalamic nuclei have previously been identified. Preliminary studies in our lab have: (1) identified serotonergic neurons responsive to leptin in the DRN that project to the arcuate nucleus (ARC) of the hypothalamus and (2) demonstrated leptin injected into the DRN significantly decreases food intake. The objective of the current study was to identify the role of 5-HT in leptin’s regulation of food intake first within the DRN, then between the DRN and the ARC. Adult male Sprague Dawley rats underwent stereotaxic surgery for guide cannula implantation in the DRN. After recovery, animals were administered 100 µg of p-chlorophenylalanine (PCPA), an inhibitor of 5-HT synthesis, in the DRN each day for four days. On the fourth day, leptin was also administered in the DRN (5 µg/rat) and food intake was measured over a 24-hour time course. ANOVA analysis revealed a significant difference in 24-hour food intake [F (3, 18) = 3.972; P = 0.0246] and post-hoc analysis showed that animals treated with leptin significantly decreased food intake (17.2 ± 2.0 g) compared to control rats (25.4 ± 0.9 g), whereas PCPA-treated rats did not differ from the control rats, suggesting that depletion of 5-HT attenuated leptin’s ability to regulate food intake within the DRN. To examine the role of 5-HT on leptin’s hypothalamic action, a subsequent experiment was conducted by implanting an additional cannula into the ARC for the administration of leptin or vehicle on the fourth day of treatment. ANOVA analysis revealed a significant difference in 24-hour food intake [F (3, 16) = 5.998; P = 0.0061] and post-hoc analysis showed that only rats treated with leptin in the ARC significantly decreased food intake (14.0 ± 1.5 g) compared to controls (21.8 ± 0.5 g). 5-HT depletion was assessed post-mortem using immunohistochemistry and was later quantified. Collectively, these results demonstrate that leptin’s ability to regulate food intake is dependent on 5-HT, regardless of the area of regulation (i.e. DRN or the hypothalamus).

Differential modulation of the anterior cingulate and insular cortices on anxiogenic-like responses induced by empathy for pain

Mice cohabiting with a conspecific in chronic pain display anxiogenesis in the elevated plus-maze (EPM). Given that the anterior cingulate (ACC) and insular (InC) cortices play a role in the modulation of anxiety, pain, and emotional contagion, we investigated (a) the FosB activation in both brain areas and (b) the effects of intra-ACC or -InC injection of cobalt chloride (CoCl2, a synaptic blocker), on the anxiety of mice cohabiting with a cagemate suffering pain. Twenty-one days after birth, male Swiss mice were housed in pairs for 14 days to establish familiarity. On the 14th day, mice were divided into two groups: cagemate sciatic nerve constriction (CNC; i.e., one animal of each pair was subjected to sciatic nerve constriction), and cagemate sham (CS; i.e., a similar procedure but without suffering nerve constriction). After that, both groups were housed again with the same pairs for the other 14 days. On the 28th day, mice had their brains removed for the immunoassays analyses (Exp. 1). For experiments 2 and 3, on the 23rd day, the cagemates received guide cannula implantation bilaterally in the ACC or InC and, on the 28th day, they received local injections of saline or CoCl2, and then were exposed to the EPM. Results showed that cohabitation with a conspecific with chronic pain decreases and increases neuronal activation (FosB) within the ACC and InC, respectively. Intra-ACC or InC injection of CoCl2 reversed the anxiogenic effect in those animals that cohabited with a conspecific in chronic pain. ACC and InC seem to modulate anxiety induced by emotional contagion in animals cohabitating with a conspecific suffering pain.

Stereotaxic surgery for implantation of guide cannulas for microinjection into the dorsomedial hypothalamus in young rats

Stereotaxic surgery to implant guide cannulas into the rodent brain is a frequently used technique to deliver drugs to targeted brain regions in awake, freely moving animals. There are limited reports, however, of central injections in young animals, and no information on cannula implantation for drug administration into the dorsomedial hypothalamus (DMH) in young rats. Our protocol describes a simple and successful method for implanting guide cannulas in the brains of young, male Sprague-Dawley rats and outlines newly developed stereotaxic coordinates to accurately target the dorsomedial hypothalamus.•Stereotaxic surgical procedure for guide cannula implantation in the DMH in young rats.•Development of stereotaxic coordinates of the DMH in young rats.•Microinjection of drugs into the young rat brain.

Theoretical and practical applications of the intracerebroventricular route for CSF sampling and drug administration in CNS drug discovery research: A mini review

Clinically, central nervous system (CNS) disorders account for more hospitalisations and prolonged care than almost all other diseases combined. In the preclinical setting, the intracerebroventricular (ICV) route for cerebrospinal fluid (CSF) sampling or dose administration in rodent models of human CNS disorders has potential to provide key insight on the pathobiology of these conditions. Low level neuroinflammation is present in >40% of patients with severe depression or schizophrenia and so comparative assessment of CSF composition between patients and rodent models of CNS disorders is potentially invaluable for hypothesis generation and for assessing rodent model validity. As molecules in the CSF have relatively low protein binding and are freely exchanged into the extracellular fluid of the brain parenchyma, supraspinal drug administration into the CSF can produce therapeutic drug concentrations in the brain. Direct administration of investigational agents into the CSF of the lateral ventricle of the brain enables intrinsic efficacy and adverse effect profiles to be evaluated without the confounding effects of drug metabolism, due to the low capacity of the CNS to metabolise exogenous compounds. It is our view that the ICV route for CSF sampling and for administration of novel drugs in development is under-utilised in preclinical research on CNS disorders. This is due to the high degree of technical skill and low margin for error associated with correct ICV guide cannula implantation in the rat. However, these technical challenges can be overcome by using standardised procedures and attention to detail during surgery and in the post-operative period.

S23-02 Neural Correlates of Empathy

Mice cohabiting with a conspecific in chronic pain display anxiogenesis in the elevated plus-maze (EPM). Given that the anterior cingulate (ACC) and insular (InC) cortices play a role in the modulation of anxiety, pain, and emotional contagion, we investigated (a) the FosB activation in both brain areas and (b) the effects of intra-ACC or -InC injection of cobalt chloride (CoCl2, a synaptic blocker), on the anxiety of mice cohabiting with a cagemate suffering pain. Twenty-one days after birth, male Swiss mice were housed in pairs for 14 days to establish familiarity. On the 14th day, mice were divided into two groups: cagemate sciatic nerve constriction (CNC; i.e., one animal of each pair was subjected to sciatic nerve constriction), and cagemate sham (CS; i.e., a similar procedure but without suffering nerve constriction). After that, both groups were housed again with the same pairs for the other 14 days. On the 28th day, mice had their brains removed for the immunoassays analyses (Exp. 1). For experiments 2 and 3, on the 23rd day, the cagemates received guide cannula implantation bilaterally in the ACC or InC and, on the 28th day, they received local injections of saline or CoCl2, and then were exposed to the EPM. Results showed that cohabitation with a conspecific with chronic pain decreases and increases neuronal activation (FosB) within the ACC and InC, respectively. Intra-ACC or InC injection of CoCl2 reversed the anxiogenic effect in those animals that cohabited with a conspecific in chronic pain. ACC and InC seem to modulate anxiety induced by emotional contagion in animals cohabitating with a conspecific suffering pain.

Effect of surgery for cannula implantation into the hypothalamus on the behaviour and neural activity of broiler chicks

1. The effects of a surgical operation, to implant a guide cannula in the chick hypothalamus for microdialysis, on behavioural responses and neural activity in broiler chicks are described. 2. General behavioural activities (feeding, preening, sitting, drinking, cage pecking and beak wiping), open field and locomotor activity tests were conducted to evaluate the effects related to surgery in the immediate 4 d following this procedure. Perfusion of Ringer solution with high K+ after 4 d of guide cannula implantation was used to estimate the neural activity resulting from surgery through stimulation of monoamine release by in vivo brain dialysis. 3. The results of direct behavioral observations indicated that the stress provoked by surgical guide cannula insertion caused behavioural alterations that are particularly evident in the immediate days following this procedure. Open-field tests on day 4 after surgery showed that, compared to the intact control chickens, the treated chicks had a shorter latency to ambulate and defecate, with more vocalisation. Locomotor activity was less in the treated chicks than inthe controls. 4. After 4 d of guide cannula implantation, the serotonin concentration started to increase 30 min after the onset of perfusing high-K+ Ringer solution. It reached its highest value at one hour, suggesting that the 4 d after surgery is enough to alleviate some neurochemical dysfunction resulting from surgery. The results of behavioural observations, open-field and locomotor activity tests indicate that the surgical operation caused stress and fear in chicks which persisted up to 4 d.

Hippocampal acetylcholine efflux increases during negative patterning and elemental discrimination in rats

The purpose of this paper is to examine whether hippocampal acetylcholine (ACh) efflux increases during negative patterning (NP) discrimination tasks. For these tasks, a rat's response was rewarded when either a single stimulus A (tone) or stimulus B (light) was presented, but was not rewarded when the compound stimulus AB (tone + light) was presented to the NP group of rats. An elemental discrimination (E) task was given to another group (E group). In the E group, the rat's response was rewarded when one of two stimuli (e.g., tone) was presented, but not rewarded when the other stimulus (e.g., light) was presented. After reaching a learning criterion, a guide cannula was implanted into dorsal hippocampus under anesthesia. In test sessions, rats were given the same task as before the guide cannula implantation, and ACh efflux was measured. Hippocampal ACh efflux increased during both NP and E tasks. In addition, the magnitude of increase was higher in the NP group than in the E group. Thus, over all our results demonstrate that task difficulty is a critical factor that relates to the difference in ACh efflux in the hippocampus.

Measurement of Blood–Brain Barrier Permeability of Rats with Alpha-Aminoisobutyric Acid During Microdialysis: Possible Application to Behavioral Studies

The purpose of this study was to determine the suitability of α-aminoisobutyric acid (AIB) as a marker of the integrity of the blood–brain barrier (BBB) during studies in which amino acid content in dialysates collected by microdialysis probes and behavior of the rat are studied concurrently. AIB (200 mg/kg) was injected intraperitoneally at 1800 h into 24-h fasted, ketamine–acepromazine anesthetized rats on either 10 or 30 days following guide cannula implantation. Dialysates from the medial preoptic area and blood from the tail vein were collected 1 h before and after the AIB injection. Analysis of amino acids, including AIB, in the collections was conducted by reverse-phase HPLC. In 21 of 24 rats, AIB in dialysates averaged less than 3% of plasma at 10, 30, and 50 min after AIB injection. In those rats unidirectional blood-to-brain transfer constant, K i, of AIB was constant and a good relationship was found between dialysate amino acid concentrations and those predicted from calculations of transport based on the brain uptake index (BUI) for some amino acids. AIB concentration in dialysates was greater than 10% of plasma at 30 min in only 3 of 24 rats. We conclude that AIB can be used as a marker to monitor the integrity of the BBB during serial measurements of dialysate concentrations of amino acids and has application in behavioral studies.

Involvement of the midbrain periaqueductal gray 5-HT 1A receptors in social conflict induced analgesia in mice

Recent results from our laboratory have shown that 30-bites social conflict in mice produces a high-intensity, short-term analgesia which is attenuated by systemically injected 5-HT 1A receptor agonists, such as BAY R 1531 (6-methoxy-4-(di- n-propylamino)-1,3,4,5-tetrahydrobenz( c, d)indole hydrochloride) and gepirone. The present study investigated the effects of these drugs, as well as the 5-HT 1A receptor antagonist WAY 100135 ( N- tert-butyl-3-(4-(2-methoxyphenyl)piperazine-1-yl)-2-phenylpropanamide) injected into the midbrain periaqueductal gray matter of mice on 30-bites analgesia. Four to five days after guide-cannula implantation, each mouse received microinjection of gepirone (30 nmol/0.2 μl), BAY R 1531 (10 nmol/0.2 μl), WAY 100135 (10 nmol/0.2 μl), saline (0.9% NaCl) or vehicle (saline+4% Tween 80) 5 min before either an aggressive (30 bites) or a non-aggressive interaction. Nociception was assessed by the tail-flick test made before as well as 1, 5, 10 and 20 min after social interaction. The full 5-HT 1A receptor agonist BAY R 1531 blocked, whereas, WAY 100135 and gepirone intensified 30-bites analgesia. Neither non-aggressive interaction, per se, nor the three compounds given after this type of social interaction significantly changed nociception. These results indicate that 5-HT 1A receptors in the periaqueductal gray inhibit analgesia induced by social conflict in mice.

Augmentation of the neurochemical effects of cocaine in the ventral striatum and medial prefrontal cortex following preexposure to amphetamine, but not nicotine: An in vivo microdialysis study

This study assessed the ability of cocaine to increase synaptic levels of dopamine (DA) in the ventral striatum (VS) and medial prefrontal cortex (mPFC) following repeated daily exposure to amphetamine or nicotine. In vivo microdialysis was used to assay DA levels in the awake freely moving male Sprague-Dawley rat. Three days following guide cannula implantation, subjects received 9 daily preexposure injections of amphetamine (1.0 mg/kg salt weight, IP), nicotine (0.6 mg/kg base weight, SC), or vehicle. Probes were then inserted and perfused with Ringer's solution overnight. Once stable baseline levels of DA were established, the subjects received a cocaine (15 mg/kg, IP) challenge injection and dialysate samples were collected every 20 minutes. Subjects preexposed to vehicle exhibited a 200% increase in DA levels in the VS and a 170% increase in the mPFC following the cocaine injection. DA recovery was significantly augmented in both regions in amphetamine-preexposed subjects with 450% and 285% increases above baseline levels observed in the VS and mPFC, respectively. In contrast, cocaine-induced DA levels in nicotine-preexposed subjects did not differ significantly from vehicle-preexposed controls in either brain region. Data are discussed in terms of alterations in the locomotor activating and reinforcing effects of cocaine following similar preexposure procedures.

Acetylcholine measurement of cerebrospinal fluid by in vivo microdialysis in freely moving rats.

Acetylcholine (ACh) and choline (Ch) levels in rat cerebrospinal fluid (CSF) were determined by in vivo microdialysis (CSF microdialysis) in both halothane-anesthetized and freely-moving rats. The Ch/ACh ratio in CSF perfused with Ringer's solution (30 microliters/30 min) containing 10(-5) M physostigmine, a centrally active cholinesterase inhibitor, was significantly lower than that in unprocessed CSF due to significantly higher ACh levels in the former. The successive measurement on the 2nd and 7th day after the guide cannula implantation demonstrated the feasibility of the CSF microdialysis method for repetitive monitoring of CSF ACh and Ch levels in freely moving rats without extensive tissue damage. Intraperitoneal administration of physostigmine caused an increase in CSF ACh levels, whereas administration of neostigmine, which cannot penetrate into the blood brain barrier, did not. Furthermore, a centrally active acetylcholinergic M1-receptor agonist, AF102B, produced an increase in CSF ACh and Ch levels. Thus, the present study demonstrates that CSF microdialysis is a useful method for evaluating overall central cholinergic activity and investigating the pharmacological effects of various drugs that act via the central cholinergic system.