Immunolocalization of Metabotropic Glutamate Receptor 1 (mGluR1) in Distinct Classes of Interneuron in the CA1 Region of the Rat Hippocampus Francesco Ferraguti, 1 * Philip Cobden, 1 Marie Pollard, 1 David Cope, 1 Ryuichi Shigemoto, 2 Masahiko Watanabe, 3 and Peter Somogyi 1 ABSTRACT: In the hippocampal CA1 region, metabotropic glutamate subtype 1 (mGluR1) receptors have been implicated in a variety of phys- iological responses to glutamate, which include modulation of synaptic transmission and plasticity, as well as neuronal excitability and synchro- nization. The mGluR1 isoform is characteristically expressed only by nonprincipal cells, and it is particularly enriched in somatostatin (SS)- containing interneurons in stratum oriens-alveus. Anatomical and physi- ological data have indicated the presence of mGluR1 in several distinct classes of interneurons with their somata located also in strata pyrami- dale, radiatum, and lacunosum moleculare. Each different interneuron subtype, as defined by functionally relevant criteria, including input/ output characteristics and expression of selective molecular markers, subserves distinct functions in local hippocampal circuits. We have inves- tigated which of the different CA1 interneuron classes express mGluR1 by immunofluorescent labeling, combining antibodies to mGluR1, calcium- binding proteins, and neuropeptides, and by intracellular labeling in vitro. Several types of interneuron that are immunopositive for mGluR1 each targeted different domains of pyramidal cells and included (1) O-LM inter- neurons, found to coexpress both SS and parvalbumin (PV); (2) interneurons with target selectivity for other interneurons, expressing vasoactive intestinal polypeptide (VIP) and/or the calcium-binding protein calretinin; (3) pro- cholecystokinin-immunopositive interneurons probably non-basket and dendrite-targeting; and (4) an as-yet unidentified SS-immunoreactive but PV-immunonegative interneuron class, possibly corresponding to oriens- bistratified cells. Estimation of the relative proportion of mGluR1-posi- tive interneurons showed 43%, 46%, and 30% co-labeling with SS, VIP, or PV, respectively. The identification of the specific subclasses of CA1 interneurons expressing mGluR1 provides the network basis for assess- ing the contribution of this receptor to the excitability of the hippocam- pus. © 2004 Wiley-Liss, Inc. KEY WORDS: immunohistochemistry; cholecystokinin; parvalbumin; mGluR7; somatostatin; vasoactive intestinal polypeptide INTRODUCTION Local -aminobutyric acid (GABA)ergic interneurons play multiple roles in the control of the output and network activity patterns of principal cells in the hippocampus (Traub and Miles, 1991; McBain and Fisahn, 2001). Physiological and anatomical evidence points to the existence of several distinct classes of inter- neurons, each potentially serving different functions in hippocampal circuits (Ramon y Cajal, 1893; Lorente de No, 1934; Freund and Buzsaki, 1996; Somogyi et al., 1998). Different classes of interneuron selectively inner- vate distinct membrane domains of pyramidal cells (So- mogyi et al., 1998) or are specialized to control other interneurons (Freund and Buzsaki, 1996). The selective expression of neurochemical markers, such as calcium- binding proteins, neuropeptides, and neurotransmitter receptors, by subpopulations of interneurons has facili- tated their classification into different populations (Kawaguchi et al., 1987; Nitsch et al., 1990; Freund and Buzsaki, 1996; Somogyi et al., 1998). In the hippocampus, one of the alternatively spliced isoforms of the metabotropic glutamate receptor subtype 1 (i.e., mGluR1), is characteristically expressed only by nonprincipal cells. It is particularly enriched in soma- tostatin (SS)-containing interneurons of the CA1 stra- tum oriens-alveus (O/A) (Baude et al., 1993; Hampson et al., 1994). Scattered interneurons immunopositive for mGluR1 have also been reported in strata pyramidale, radiatum, and lacunosum moleculare (Martin et al., 1992; Baude et al., 1993; Lujan et al., 1996; Shigemoto et al., 1997). Alternative splicing at the mGluR1 gene generates at least two other receptor proteins: mGluR1 and mGluR1d (Tanabe et al., 1992; Laurie et al., 1996). These splice variants are expressed in the hippocampus in both principal and nonprincipal cells (Shigemoto et al., 1992; Berthele et al., 1998; Ferraguti et al., 1998). Im- munoreactivity (IR) for mGluR1 is particularly en- riched in interneurons and pyramidal cells of the CA3 area but was undetectable in the CA1 area (Ferraguti et al., 1998). The location of the mGluR1d receptor pro- tein is unknown, because antibodies that selectively iden- tify this variant are not yet available. Nevertheless, some interneurons in strata oriens and pyramidale of the CA1 area were shown to have abundant mRNA expression for mGluR1d (Berthele et al., 1998). A variety of physiological responses to glutamate ap- pear to implicate mGluR1 receptors in the CA1 area. These responses include the modulation of synaptic transmission and plasticity (McBain et al., 1994; 1 MRC Anatomical Neuropharmacology Unit, Department of Pharmacol- ogy, Oxford University, Oxford, United Kingdom; 2 Division of Cerebral Structure, National Institute for Physiological Sciences, Myodaiji, Oka- zaki, Japan; 3 Department of Anatomy, Hokkaido University School of Medicine, Sapporo, Japan *Correspondence to: Francesco Ferraguti, MRC Anatomical Neuropharma- cology Unit, Department of Pharmacology, Oxford University, Mansfield Road OX1 3TH, Oxford, UK. E-mail: francesco.ferraguti@pharm.ox.ac.uk Accepted for publication 5 March 2003 DOI 10.1002/hipo.10163 HIPPOCAMPUS 14:193–215 (2004) © 2004 WILEY-LISS, INC.