Expression of the pacemaker channel HCN4 in excitatory interneurons in the dorsal horn of the murine spinal cord.

In the central nervous system, hyperpolarization-activated, cyclic nucleotide-gated (HCN1-4) channels have been implicated in neuronal excitability and synaptic transmission. It has been reported that HCN channels are expressed in the spinal cord, but knowledge about their physiological roles, as well as their distribution profiles, appear to be limited. We generated a transgenic mouse in which the expression of HCN4 can be reversibly knocked down using a genetic tetracycline-dependent switch and conducted genetically validated immunohistochemistry for HCN4. We found that the somata of HCN4-immunoreactive (IR) cells were largely restricted to the ventral part of the inner lamina II and lamina III. Many of these cells were either parvalbumin- or protein kinase Cγ (PKCγ)-IR. By using two different mouse strains in which reporters are expressed only in inhibitory neurons, we determined that the vast majority of HCN4-IR cells were excitatory neurons. Mechanical and thermal noxious stimulation did not induce c-Fos expression in HCN4-IR cells. PKCγ-neurons in this area are known to play a pivotal role in the polysynaptic pathway between tactile afferents and nociceptive projection cells that contributes to tactile allodynia. Therefore, pharmacological and/or genetic manipulations of HCN4-expressing neurons may provide a novel therapeutic strategy for the pain relief of tactile allodynia.

Evidence of specialized tissue in human interatrial septum: histological, immunohistochemical and ultrastructural findings.

BACKGROUND: There is a paucity of information on structural organization of muscular bundles in the interatrial septum (IAS). The aim was to investigate histologic and ultrastructural organization of muscular bundles in human IAS, including fossa ovalis (FO) and flap valve. METHODS: Macroscopic and light microscopy evaluations of IAS were performed from postmortem studies of 40 patients. Twenty three IAS specimens underwent serial transverse sectioning, and 17--longitudinal sectioning. The transverse sections from 10 patients were immunolabeled for HCN4, Caveolin3 and Connexin43. IAS specimens from 6 other patients underwent electron microscopy. RESULTS: In all IAS specimens sections the FO, its rims and the flap valve had muscle fibers consisting of working cardiac myocytes. Besides the typical cardiomyocytes there were unusual cells: tortuous and horseshoe-shaped intertangled myocytes, small and large rounded myocytes with pale cytoplasm. The cells were aggregated in a definite structure in 38 (95%) cases, which was surrounded by fibro-fatty tissue. The height of the structure on transverse sections positively correlated with age (P = 0.03) and AF history (P = 0.045). Immunohistochemistry showed positive staining of the cells for HCN4 and Caveolin3. Electron microscopy identified cells with characteristics similar to electrical conduction cells. CONCLUSIONS: Specialized conduction cells in human IAS have been identified, specifically in the FO and its flap valve. The cells are aggregated in a structure, which is surrounded by fibrous and fatty tissue. Further investigations are warranted to explore electrophysiological characteristics of this structure.

Myocardial expression of hyperpolarization-activated, cyclic nucleotide-gated proteins in healthy cats and cats with hypertrophic cardiomyopathy.

In this article the myocardial expression of different hyperpolarization-activated, cyclic nucleotide-gated (HCN) isoforms in myocardial tissue from healthy control cats and cats with hypertrophic cardiomyopathy (HCM) was evaluated. Myocardial tissue samples of the left ventricle of control cats (n = 12) and cats with HCM (n = 4) were collected. Expression of feline HCN was determined by immunoblot analysis using antibodies against HCN2 and HCN4. Optical densities of HCN bands were compared among groups by use of the Mann-Whitney Rank Sum test. HCN4 was reliably detected in myocardial tissue whereas HCN2 was not. HCN4 expression was significantly increased in left ventricular (LV) myocardial samples of cats with HCM (P = 0.036) compared to control cats. Results indicate that myocardial HCN4 expression can be evaluated in cats by immunoblot analysis and that HCN4 expression is upregulated in LV myocardial tissue of cats with HCM. The pathophysiological importance of HCN overexpression with regard to myocyte function and altered automaticity deserves further study.