Journal of Neuroscience Methods, 50 (1993) 237-241 237 © 1993 Elsevier Science Publishers B.V. All rights reserved 0165-0270/93/$06.00 NSM 01570 A remote insertion technique for intracerebral microinjections in freely moving animals Marco A. Parada a,b,,, Marina Puig de Parada a,b and Bartley G. Hoebel b a Los Andes University, School of Medicine, M~rida (Venezuela) and b Department of Psychology, Princeton University, Princeton, NJ (USA) (Received 11 December 1992) (Revised version received 11 June 1993) (Accepted 14 June 1993) Key words: Microinjection; Cannula; Technique; Nervous system; Brain This report describes two improvements to the typical double-cannula microinjection technique. (1) Intracerebral microinjec- tions usually require holding the animal during the insertion of an injector through an implanted guide cannula; however, this is not necessary with the technique described. The injector is made of a long piece of fused silica capillary tubing (145 mm outer diameter x 21.2 cm) which is so small and flexible that it slips through a PE-20 tube (20 cm) that guides it into the implanted guide cannula and down to the desired brain site where it stops. (2) Connection to a microliter syringe is usually done with PE tubing which is leaky, expandable and represents a relatively large dead space that makes it difficult to deliver small, accurate volumes. This problem is avoided by making connection to the syringe via another piece of silica glass capillary tubing. Thus both the injector and its connection to the syringe are made of glass. With these modifications the injector can be inserted without touching the animal, and accurate volumes in the low nanoliter range can be delivered. Direct administration of substances into cir- cumscribed regions of the brain in awake animals has been used in physiological psychology and behavioral neuroscience research for at least 40 years (Anderson, 1953; Grossman, 1960). Mi- croinjections are useful for localized application of various blood-borne substances such as inter- leukin (Plata-Salam~in and ffrench-Mullen, 1992), extracellular constituents such as sodium chloride (Anderson, 1953), neurotransmitters such as norepinephrine (Grossman, 1960; Leibowitz, 1978), drugs, neurotransmitters plus drugs or neurotransmitters compared to drugs (Leibowitz, 1975; Leibowitz and Rossakis, 1979; Parada et al., 1988, 1992), radiographic or fluorescent tracers (Phillipson, 1978), neurotoxins (Ungerstedt, 1971; * Corresponding author: Dr. Marco A. Parada, Departamento de Fisiologla, Apartado Postal 109, M6rida 5101-A, Venezuela. Tel.: 11-58-74-403111; FAX: 11-58-74-634587. Ahlskog et al., 1984), peptides (Stanley et al., 1985) and hormones (Sakaguchi and Bray, 1988) to name a few. This report describes two modifi- cations to traditional techniques (Hoebel, 1964; Myers, 1971) that render the microinjection an easier and more effective tool in neuroscience research. The first modification allows insertion of an injector without touching the animal by using a foot-long injector of glass tubing, and the second increases the accuracy of injecting nano- liter volumes into the brain by connecting the injector to the remote syringe with another piece of glass tubing. This technique for inserting an injector with- out touching the animal requires construction of 4 components: an intracerebral guide cannula, its obturator stylet, an injector, and the external guide for the remote insertion of the injector into the intracerebral guide (Fig. 1). The intracerebral guide cannula (Fig. 1A and D) is made of 2 segments of stainless steel tubing