Osmotic Minipump Implantation for Increasing Glucose Concentration in Mouse Cerebrospinal Fluid

Abstract

Diabetes increases the risk of cognitive decline and impairs brain function. Whether or not this relationship between high glucose and cognitive deficits is causal remains elusive. Moreover, whether these deficits are mediated by an increase in glucose levels in cerebrospinal fluid (CSF) and/or blood is also unclear. There are very few studies investigating the direct effects of high CSF glucose levels on central nervous system (CNS) function, especially on learning and memory, since current diabetes models are not sufficiently developed to address such research questions. This article describes a method to chronically increase CSF glucose levels for 4 weeks by continuously infusing glucose into the lateral ventricle using osmotic minipumps in mice. The protocol was validated by measuring glucose levels in CSF. This protocol increased CSF glucose levels to ~328 mg/dL after infusion of a 50% glucose solution at a 0.25 µL/h flow rate, compared to a CSF glucose concentration of ~56 mg/dL in mice that received artificial cerebrospinal fluid (aCSF). Furthermore, this protocol did not affect blood glucose levels. Therefore, this method can be used to determine the direct effects of high CSF glucose on brain function or a specific neural pathway independently of changes in blood glucose levels. Overall, the approach described here will facilitate the development of animal models for testing the role of high CSF glucose in mediating features of Alzheimer's disease and/or other neurodegenerative disorders associated with diabetes.

Figure 1:. Assembly of osmotic minipumps.

(A) Infusion assembly with a cannula connected to a minipump through tubing. These pumps require at least 48 h to prime. (B) The presence of air bubbles outside the minipumps confirms the priming.

Figure 2:. Stereotaxic apparatus and accessories.

(A,B) Stereotaxic equipment with an attached micromanipulator and other accessories. (C) Burr hole coordinates to insert the cannula. (D) Osmotic mini pump implantation, (E,F) Insertion of the cannula in the drilled hole. Maintain aseptic conditions throughout the surgery.

Figure 3:. Cerebrospinal fluid (CSF) collection procedure.

(A) The dorsal neck muscles were gently displaced with blunt forceps to expose the cisterna magna. A 1 mm capillary with a 0.5 mm diameter tip was used to (B) rupture and (C,D) collect CSF from the cisterna magna.

Figure 4:. Measurement of glucose.

(A) Increased CSF glucose (B) without affecting non-fasting blood glucose levels in mice infused with 50% glucose solution in the lateral ventricle. The efficacy of this protocol was validated by measuring CSF and blood glucose concentration 10 days after initiating the glucose infusion. Mice infused with 50% glucose solution had CSF glucose levels of 327.7 ± 30.1 mg/dL (mean ± standard error of mean) compared to mice that received artificial CSF infusion that had glucose levels of 56.5 ± 2.6 mg/dL. ****p < 0.0001, unpaired t-test. Error bars represent standard error of mean (n = 5).